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  • 1985-1989  (9,226)
  • 1980-1984  (5,447)
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  • 101
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 264-272 
    ISSN: 0886-1544
    Keywords: rotary shadowing ; microtubules ; cytoplasmic movement ; conformation change ; two-headed molecule ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The molecular structure of bovine adrenal kinesin was studied by electron microscopy using the low-angle rotary shadowing technique. Adrenal kinesin exhibited either a folded or an extended configuration; the ratio of the two is dependent on the salt concentration. Almost all adrenal kinesin molecules were folded in a low-ionic solution, and the ratio of extended molecules increased to 40-50% in a solution containing 1 M ammonium acetate. Kinesin in the extended configuration displayed a rod-shaped structure with a mean length of about 80 nm. The morphologies of the ends were different; one end was composed of two globular particles, similar to the two-headed structure of myosin, while the other end had a more ill-defined structure, appearing either as a globular particle, an aggregate of two to four small granules, or a frayed, fan-like structure. The folded kinesin molecule possessed a hinge region in the middle of the rod, at about 32 nm from the neck of the two heads. In our preparations, the majority of adrenal kinesin molecules were folded at physiological salt concentrations. Adrenal kinesin bound to microtubules in the presence of adenylyl imidodiphosphate (AMP-PNP) also displayed a folded morphology.
    Additional Material: 7 Ill.
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  • 102
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 103
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 345-358 
    ISSN: 0886-1544
    Keywords: cell motility ; microtubules ; mussel gill ; ATPase ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In the presence of specific inhibitors of beat, 20 μM VO43- or pCa 4, mussel gill lateral (L) cilia can be arrested in two positions - “hands down” or “hands up” - at opposite ends of the stroke cycle. Cilia move to these positions by doublet microtubule sliding. Axonemes of arrested cilia, still tethered to the cell, are intact after demembranation and protease treatment. When reactivated by 4 mM ATP with inhibitors present, about 40% split apart. Splits are not random but occur preferentially between different specific doublets in the two opposite arrest positions. Several different related patterns of splitting are observed; for every pattern in “hands down” axonemes, there is a corresponding complementary split pattern in “hands up” axonemes. In some split patterns two doublets remain firmly attached to the central pair; these also differ depending on axonemal position. Although some of the patterns seen may be artifactual or difficult to explain, the complementary splitting patterns are predictable with simple assumptions by a “switch point” hypothesis of ciliary activity where, during each recovery stroke, doublets 6-8 have active dynein arms, while during each effective stroke, arms on doublets 1-4 become active, and arms 6-8 are turned off. Because of a difference between the patterns seen and the predictions, the status of the arms on doublet 9 is unresolved. The patterns also suggest that a spokecentral sheath attachment cycle may correlate with switching of arm activity during the generation of an asymmetric beat.
    Additional Material: 8 Ill.
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  • 104
    ISSN: 0886-1544
    Keywords: microtubule ; membrane organelle ; cross bridges ; intracellular motion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two major brain microtubule-associated proteins (MAPs), MAP2 and tau, were found to be able to bind to purified rat brain mitochondria. The apparent dissociation constants of the binding of thermostable 32P-labeled MAP2 and tau are 0.9 ± 0.04 × 10-7 and 3.8 ± 0.7 × 10-7 M, respectively. 32P-labeled MAP2 and tau bound to the mitochondria can be displaced by phosphorylated, nonradioactive MAP2. The binding parameters of MAP2 prepared without heat treatment and those of the thermostable MAP2 were of the same order of magnitude. Microtubule-binding and projection domains of MAP2 were obtained by chymotryptic digestion of rat brain microtubules (Vallee, Proc. Natl. Acad. Sci. USA, 77:3206-3210, 1980). Displacement studies with these two domains show that MAP2 bound to mitochondria can be displaced by the microtubule-binding domain, whereas the projection domain does not displace MAP2. The two domains of MAP2 bind to the mitochondria with similar affinity constants; however, the Bmax for the projection domain was 10 times and 35 times lower than the Bmax of the binding of the intact MAP2 and the microtubule-binding domain, respectively. Chymotryptic digestion of MAP2 bound to the mitochondria yielded peptide fragments with molecular masses similar to those obtained by the digestion of MAP2 bound to the microtubules. The fragments corresponding to the projection domain were released into the extramitochondrial supernatant, whereas the fragments originating from the microtubule-binding domain remained bound to the mitochondria. These results suggest that MAP2 binds to mitochondria preferentially via its microtubule-binding domain.
    Additional Material: 6 Ill.
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  • 105
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 359-371 
    ISSN: 0886-1544
    Keywords: chymotrypsin digest ; multiple immunoblot ; keratin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Affinity-purified antibodies raised against three flagellar tektins (tektin A, B, and C) from each of two sea urchin species (Lytechinus pictus and Strongylocentrotus purpuratus) were used to study the immunological relationship between tektins and intermediate filament proteins. By immunofluorescence microscopy, several antitektins revealed a staining of intermediate filament-like arrays in three vertebrate cell lines tested. Immunoelectron microscopy substantiated the cross reaction of antitektins with intermediate filaments. When the cells were treated with cytochalasin B, the arrangement of the filaments recognized by anti-(Lp)-tektin B was altered; the alteration observed is typical for keratin filaments. By immunoblot, it was found that anti-(Lp)-tektin B cross reacted with two isoforms or different proteins of ∼54 kD with pIs of 6.1 and 6.2 in human carcinoma epithelia (HeLa) cells and with two isoforms or different proteins of ∼55 kD with pIs of 6.1 and 6.3 in pig kidney epithelia (LLC-PK1) cells. Furthermore, when antitektin antibodies were affinity purified with the 54 kD HeLa keratin, these keratin-specific antibodies again restained the original tektins on immunoblots. From these observations, it can be concluded that tektins and keratins are to a certain extent immunologically related. To determine the degree of the immunological relationship, tektin filaments and purified intermediate filaments from HeLa cells were cleaved with α-chymotrypsin and examined by quantitative immunoblot analysis. On immunoblots of digested tektins from L. pictus, anti-(Lp)-tektin B recognized several cleavage products in the range of 20 kD to 46 kD. However, when immunoblots of digested intermediate filaments from HeLa cells were probed, the cross reaction of anti-(Lp)-tektin B with HeLa keratins was eliminated by more than 98% within 2 min, suggesting that tektins have epitopes in common with the end domains of certain keratins.
    Additional Material: 8 Ill.
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  • 106
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 382-392 
    ISSN: 0886-1544
    Keywords: motility ; cell surface ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have used 400 kilovoit intermediate voltage electron microscopy (IVEM) to examine thick sections of fibroblasts cultured in collagen gels. In these 3D collagen lattices, the long, narrow pseudopodial extensions that extend out and make contact with the collagen matrix exhibit a complex topography not seen in the processes put out by cells moving on planar substrata. For this reason, sections 1 to 2 μm thick that enclose a whole cell process are more informative of the overall morphology of the interaction between cells and the collagen than are thin sections. To aid the discrimination of topography of cell processes in stereo views of micrographs, some cells were labeled with antibodies and protein A-colloidal gold conjugates. The gold particles provided clear 3D reference points for computeraided reconstructions of membrane topography from tilt series of IVEM images. Our results confirm that cells that move through collagen lattices lack the wellspread morphology of their counterparts moving on glass. They are generally rather spindly with several long branching anterior pseudopodia. We found that the cell bodies and major pseudopodial processes were cylindrical, as one might expect of cells in a 3D environment, but at the leading edge of advancing pseudopodia there are small flat extensions similar to those seen in cells on glass. This similarity suggests that the lamellipodium is a basic type ofprotrusive structure used by fibroblasts during locomotion on all types of substratum. The flattened shape of lamellipodia may be part of the mechanism by which cells sense the orientation of fibrillar extracellular matrices during embryonic morphogenesis.
    Additional Material: 8 Ill.
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  • 107
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 169-180 
    ISSN: 0886-1544
    Keywords: microtubules (acetylated) ; neuronal differentiation ; map 2 ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two posttranslational modifications of alpha-tubulin, acetylation and detyrosination, are associated with stable microtubule (MT) populations, including those of neuronal processes. We have used a pluripotent embryonal carcinoma cell line, P19, to investigate changes in MT isotype and stability found in MT arrays during neurogenesis. This cell line has an advantage in that both commitment- and differentiation-related events can be observed. Uncommitted P19 cells have minimal arrays of acetylated and detyrosinated MTs. Following neuronal induction with retinoic acid (RA), indirect immunofluorescence microscopy shows that the first MT modifications occur during commitment and before any morphological change is observed. RA-induced cells initially polymerize a temporarily enlarged population of MTs. Included in this population is a new array of acetylated MTs arranged in a bundle of parallel MTs. This bundle is colchicine-stable, although no MT-associated proteins (MAPs) are detectable using a battery of anti-MAP antibodies. Observation of MT arrays with patterns that are intermediate between the early bundles and short neurites suggests that the acetylated MT bundle subsequently extends to form a neurite. MAP 2 is first detected at about the time of neurite extension. However, at this early stage of differentiation, MAP 2 is not yet limited to dendritic processes. This report provides the first evidence that the stable MTs of mature neurons may be initiated during neuronal commitment.
    Additional Material: 8 Ill.
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  • 108
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 225-247 
    ISSN: 0886-1544
    Keywords: spectrin ; ankyrin ; protein 4.1 ; membrane skeleton ; spectrin-filament interaction ; fodrin ; adducin ; calpactin I ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The purpose of this review on spectrin is to examine the functional properties of this ubiquitous family of membrane skeletal proteins. Major topics include spectrin-membrane linkages, spectrin-filament linkages, the subcellular localization of spectrins in various cell types and a discussion of major functional differences between erythroid and nonerythroid spectrins. This includes a summary of studies from our own laboratories on the functional and structural comparison of avian spectrin isoforms which are comprised of a common alpha subunit and a tissue-specific beta subunit. Consequently, the observed differences among these spectrins can be assigned to differences in the properties of the beta subunits.
    Additional Material: 4 Ill.
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  • 109
    ISSN: 0886-1544
    Keywords: latrunculin A ; latrunculin B ; cell shape ; actin organization ; cell growth and division ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The latrunculins are architecturally novel marine compounds isolated from the Red Sea sponge Latrunculia magnifica. In vivo, they alter cell shape, disrupt microfilament organization, and inhibit the microfilament-mediated processes of fertilization and early development. In vitro, latrunculin A was recently found to affect the polymerization of pure actin in a manner consistent with the formation of a 1:1 molar complex with G-actin. These in vitro effects as well as previous indications that the latrunculins are more potent than the cytochalasins suggest differences in the in vivo mode of action of the two clases of drugs. To elucidate these differences we have compared the short- and long-term effects of latrunculins on cell shape and actin organization to those of cytochalasin D. Exposure of hamster fibroblast NIL8 cells for 1-3 hr to latrunculin A, latrunculin B, and cytochalasin D causes concentration-dependent changes in cell shape and actin organization. However, the latrunculin-induced changes were strikingly different from those induced by cytochalasin D. Furthermore, while initial effects were manifest with both latrunculin A and cytochalasin D already at concentrations of about 0.03 μg/ml, latrunculin A caused complete rounding up of all cells at 0.2 μg/ml, whereas with cytochalasin D maximum contraction was reached at concentrations 10-20 times higher. The short-term effects of latrunculin B were similar to those of latrunculin A although latrunculin B was slightly less potent. All three drugs inhibited cytokinesis in synchronized cells, but their long-term effects were markedly different. NIL8 cells treated with latrunculin A maintained their altered state for extended periods. In contrast, the effects of cytochalasin D progressed with time in culture, and the latrunculin B-induced changes were transient in the continued presence of the drug. These transient effects were found to be due to a gradual inactivation of latrunculin B by serum and were used to compare recovery patterns of cell shape and actin organization in two different cell lines. This comparison showed that the transient effects of latrunculin B were fully reversible for the NIL8 cells and not for the mouse neuroblastoma N1E-115 cells.
    Additional Material: 10 Ill.
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  • 110
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989), S. 195-211 
    ISSN: 0886-1544
    Keywords: cell shape ; cortical actin ; stress fibers ; microfilament bundles ; cell adhesion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The outline of cells in sparse cultures consists prediminantly of concave and convex segments; straight segments are rare and ephemeral. The convex segments are areas of active cell expansion. The concave segments are stationary and web-shaped, similar in profile to the cables of a suspension bridge. In 3T3 fibroblasts, we have found a single microfilament bundle following the outline of every webbed edge and have called it the actin edge-bundle (AEB). While the AEB is composed predominantly of actin, α-actinin and myosin are also present. In contrast to normal stress fibers, AEBs are more resistant to several treatments that depolymerize F-actin. Once an AEB disassembles, however, the webbed edge collapses and retracts, suggesting that the actin edge-bundle is a specialized cytoskeletal structure that supports the webbed edges of interphase 3T3 fibroblasts. The stability of AEBs is independent of microtubules. We suggest that the microfilament bundles that frequently line the lateral contacts between epithelial cells in vivo may be related to the actin edge-bundle.
    Additional Material: 12 Ill.
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  • 111
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989), S. 94-103 
    ISSN: 0886-1544
    Keywords: cell adhesion ; cell motility ; near infrared light ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Interference-reflection microscopy (IRM) is the only method presently available with which to visualize cell-substratum adhesions in living tissue culture cells continuously for long periods of time without the use of fluorescent markers (Curtis: J. Cell Biol. 20:199-215, 1964; Izzard and Lochner: J. Cell Sci. 21:129-159, 1976). This method utilizes approximately 1% of the incident illumination to produce the IRM image (Verschueren: J. Cell Sci. 75:279-301, 1985) and so far has required the use of high-intensity light sources in the visible spectral range (400-800 nm). Unfortunately, visible light of this intensity and spectral range induces marked changes in the behavior and morphology of motile fibroblasts, including cessation of locomotion. In contrast, the present paper reports that continuous observations of live cells in IRM for periods of up to 8 hours are possible if the illuminating light is in the red to near-infrared range (650-950 nm) and without any observable change in normal cell morphology or behavior. In addition, we describe how the technique of Y-contrast image processing can be applied to IRM images to create a three-dimensional image of the ventral cell surface topography.
    Additional Material: 9 Ill.
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  • 112
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 113
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 104-117 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Additional Material: 5 Ill.
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  • 114
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 81-91 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Additional Material: 4 Ill.
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  • 115
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 116
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 42-52 
    ISSN: 0886-1544
    Keywords: amoeboid movement ; endocytosis ; cation composition ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: To study the in vivo role of myosin-II in Acanthamoeba castellanii, motile cells were microinjected with monoclonal antibodies raised against the myosin-II heavy chain. All injected cells underwent a transient shock response. It was found that although injection of buffer alone or of an endogenous Acanthamoeba protein decreased the motility of injected cells from 7 μm/min to ∼3 μm/min, injection of monoclonal antibodies specific for myosin-II decreased motility further to ∼0.8 μm/min. This effect was seen whether or not the monoclonal antibody to myosin-II inhibited the actomyosin-II MgATPase activity in vitro. Levels of antibody far in excess of endogenous myosin-II concentrations could not completely block amoeboid movement. The morphology of moving antimyosin-II-injected cells was unusual, suggesting a greater defect in the ability to retract the trailing edge of the cell rather than to extend the leading edge. Endosomes frequently disappeared from injected cells, and although buffer-injected cells rapidly recovered visible endosomes (50% recovery at 5 min), endosomes were not seen in antimyosin-II-injected cells until, on the average, ∼50 min after injection. Injection of a nonspecific antibody or of a nonspecific exogenous protein (ovalbumin) also decreased the mobility of the injected cells beyond that of buffer-injected cells (to ∼1 μm/min). These cells tended to recover endosomes more rapidly (∼25 min) than cells injected with antimyosin-II monoclonal antibodies. The inability of antibodies to myosin-II to inhibit completely any of the movements studied suggests that although myosin-II probably plays a role in these motilities, the cell either routinely uses or can draw upon another cytoplasmic motor to maintain locomotion, organelle movement, contractile vacuole activity, and endocytosis.
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  • 117
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 104-112 
    ISSN: 0886-1544
    Keywords: Ca2+ control ; Beroë macrocilia ; sliding disruption ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Macrocilia of the ctenophore Beroë are activated to beat continuously in the normal direction by membrane-mediated Ca2+ influx (Tamm: Journal of Comparative Physiology [A] 163:23-31, 1988a). Using saponin or Brij-58 permeabilized models of macrocilia, we show that ATP-reactivation of beating requires μM levels of free Ca2+, Ba2+, or Sr2+. Isolated macrocilia beat initially in reactivation solution (RS) containing Ca2+, Ba2+, or Sr2+ and then undergo microtubule sliding disintegration without added proteases. Addition of protease inhibitors to RS + 10-5 M Ca2+ prevents sliding disruption. Pretreatment in wash solution (containing 1 mM EGTA) without protease inhibitors, followed by RS + 10-5 M Ca2+ with protease inhibitors results in extensive sliding disintegration. However, treatment in wash solution followed by RS + protease inhibitors does not induce sliding. Therefore, Ca2+ is not required for proteolysis by endogenous proteases, but is necessary for sliding disintegration.Local iontophoretic application of Ca2+, Ba2+, or Sr2+ to permeabilized macrocilia in RS lacking these cations triggers motility and/or sliding disintegration. Extrusion of microtubules occurs from the tip or the base, depending on whether or not the macrocilium remains attached to its large actin bundle. Thin sheets of microtubules telescope out initially, due to synchronized sliding of subsets of doublet microtubules from parallel rows of axonemes.Macrocilia are one of the first examples of ATP-induced microtubule sliding which retains Ca2+ sensitivity. In addition, the finding that Ba2+ and Sr2+ also trigger active sliding provides an additional method for investigating the control of dynein-powered microtubule movements.
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  • 118
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 14 (1989), S. 237-250 
    ISSN: 0886-1544
    Keywords: mitosis ; spindle fibers ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We studied the distribution of acetylated α-tubulin in the microtubules of spermatogenic cells from the crane fly Nephrotoma suturalis (Loew) using a mono-clonal antibody specific for acetylated α-tubulin (6-11B-1). We found that cells in all stages of spermatogenesis contained acetylated microtubules including primary spermatocytes, meiotic cells, spermatids, and sperm. A subset of the acety-lated microtubules (those in midbodies and flagella) were resistant to cold depolymerization. Newly polymerized microtubules in nondividing cells were not acetylated for up to 15 min. indicating that acetylation lagged behind polymerization. In spindles, newly polymerized microtubules were acetylated after 5 min. Antibodies to acetylated α-tubulin selectively stained chromosome-to-pole fibers in dividing cells, but the staining appeared to decrease and taper of at the kinetochores. This observation supports the hypothesis that tubulin subunits add at the kinetochore in metaphase and that acetylation occurs subsequent to addition. Further, this taper may be useful as a marker in anaphase, to distinguish between different hypotheses of chromosome motion.
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  • 119
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 12 (1989), S. 248-263 
    ISSN: 0886-1544
    Keywords: ankyrin ; adducin ; protein 4.1 ; correlation length ; flexural rigidity ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The three avian spectrins that have been characterized consist of a common α-subunit (240 kD) paired with an isoform-specific β-subunit from either erythrocyte (220 or 230 kD), brain (235 kD), or intestinal brush border (260 kD). Analysis of avian spectrins, with their naturally occurring “subunit replacement” has proved useful in assessing the relative contribution of each subunit to spectrin function. In this study we have completed a survey of avian spectrin binding properties and present morphometric analysis of the relative flexibility and linearity of various avian and human spectrin isoforms. Evidence is presented that, like its mammalian counterpart, avian brain spectrin binds human erythroid ankyrin with low affinity. Cosedimentation analysis demonstrates that (1) avian erythroid protein 4.1 stimulates spectrin-actin binding of both mammalian and avian erythrocyte and brain spectrins, but not the TW 260/240 isoform, (2) calpactin I does not potentiate actin binding of either TW 260/240 or brain spectrin, and (3) erythrocyte adducin does not stimulate the interaction of TW 260/240 with actin.In addition, a morphometric analysis of rotary-shadow images of spectrin isoforms, individual subunits, and reconstituted complexes from isolated subunits was performed. This analysis revealed that the overall flexibility and linearity of a given spectrin heterodimer and tetramer is largely determined by the intrinsic rigidity and linearity of its β-spectrin subunit. No additional rigidity appears to be imparted by noncovalent associations between the subunits. The scaled flexural rigidity of the most rigid spectrin analyzed (human brain) is similar to that reported for F-actin.
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  • 120
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989), S. 9-20 
    ISSN: 0886-1544
    Keywords: carotenoid droplet ; intermediate filament ; microfilament ; microtubule ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The cytoskeleton of goldfish xanthophores contains an abundance of unique dense structures (400 nm in diameter) that are absent in goldfish nonpigment cells and are probably remnants of pterinosomes. No major difference in protein composition between xanthophores and nonpigment cells (without these structures) was found that could account for these structures. In xanthophores, these structures are foci of radiating filaments. The addition or withdrawal of ACTH causes a radical rearrangement of the xanthophore Cytoskeleton accompanying redistribution of carotenoid droplets, namely, the virtual exclusion of these dense bodies with associated filaments from the space occupied by the carotenoid droplet aggregate vs. a relatively even cytoplasmic distribution of these structures when the carotenoid droplets are dispersed. These changes in cytoskeletal morphology are not accompanied by any major changes in the protein or phosphoprotein composition of the cytoskeleton.
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  • 121
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989), S. 30-40 
    ISSN: 0886-1544
    Keywords: microtubules ; chromosome movement ; Paramecium ; nuclear lamina ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The micronuclear spindle apparatus in Paramecium bursaria was studied by electron microscopy during prophase, metaphase, and anaphase of the first meiotic division. During prophase, the spindle apparatus consists mostly of intermediate-like filaments, relatively few spindle microtubules, and unique cone-shaped structures termed microlamellae. Microlamellae join the ends of chromosomes to the fibrous elements of the spindle. The capacity to preserve the intermediate-like filaments is largely dependent upon the use of collidine buffer during fixation. In contrast, during metaphase and anaphase, microtubules are the dominant fibrous element of the spindle. The microtubules interact with chromosomes during these phases by joining to true kinetochores. Neither treatment with cytochalasin B or fixation with a low concentration of osmium tetroxide affects the development of intermediate filaments during prophase. Because intermediate-like filaments are abundant during prophase and microtubules are more common during metaphase and anaphase, the structural differences may reflect differences in the mechanisms for chromosome movement.
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  • 122
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 13 (1989) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 123
    ISSN: 0886-1544
    Keywords: kinases ; microtubules ; organelle protein ; pigment aggregate ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Triton-insoluble cytoskeleton of nonpigment cells has bound protein kinase that phosphorylates, with or without added cAMP, tubulins and the intermediate filament proteins p60, p56, p53, and p45a to give multiple charge variants. In the absence of 8-Br-cAMP, Triton-insoluble cytoskeletons from xanthophores also phosphorylate p60, p56, and p45a, but not p53; tubulin phosphorylation may also be reduced. In the presence of 8-Br-cAMP, p53, as well as several other peptides, are phosphorylated. One of these latter peptides was identified as the carotenoid droplet (pigment organelle) protein p57, whose phosphorylation and dephosphorylation precede pigment dispersion and aggregation respectively (Lynch et al.: J. Biol. Chem. 261:4204-4211, 1986). The amount of pp57 produced depends on the state of pigment distribution in the xanthophores used to prepare the cytoskeletons for labeling. With cytoskeletons from xanthophores with aggregated pigment, pp57 is a major labeled phosphoprotein seen in two-dimensional gels. With cytoskeletons prepared from xanthophores with dispersed pigment, the yield of labeled pp57 is greatly reduced (by at least 90%). Together with earlier results, we propose that, in the aggregated state, p57 serves to bind carotenoid droplets to the cytoskeletons, most likely the microtubules. The significance of other cAMP-dependent phosphorylation reactions is unknown but may be related to cAMP-induced cytoskeleton rearrangement in intact xanthophores.
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  • 124
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    Cell Motility and the Cytoskeleton 13 (1989), S. 67-82 
    ISSN: 0886-1544
    Keywords: cytoskeletal arrays ; heat shock ; synchronous CHO cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The vimentin intermediate filament (VIMF) network is more sensitive to heat-induced disruption than either the microtubule (MT) or microfilament (MF) cytoskeletal (CSK) arrays in G1 Chinese hamster ovary (CHO) cells (Coss and Wachsberger: Radiation Research, 1987). We therefore investigated the effect of the VIMF disruptive agent, acrylamide (Eckert: European Journal of Cell Biology 37:169-174, 1985), on the heat response of synchronous CHO cells. Cells, either in the process of spreading (G1 or S phase) or in the well-spread state (S phase), were exposed to a nontoxic concentration of 5 mM acrylamide, heated, and processed for immunofluorescence microscopy 30 min or 20 hr following the heat shock. Recovery from CSK disruption was related to cell survival.CHO cells, either in the process of spreading or in the well-spread state, were sensitized to heat-induced CSK disruption and cytotoxicity by acrylamide. Recovery from CSK disruption correlated with surviving fractions of cells treated in the G1 phase but not with surviving fractions of cells treated in the S phase and was independent of the degree of cell spreading. This correlation suggests that damage to CSK structures may contribute to the death of cells treated in G1 but not necessarily to the death of cells treated in S phase.The degree of acrylamide sensitization of heat-induced CSK disruption was greater for cells exposed to acrylamide prior to spreading than for well-spread cells. Furthermore, normal spreading of cells was prevented when they were plated into medium containing acrylamide, suggesting that acrylamide interferes with the initial stages of attachment and spreading of these cells. These observations are interpreted in relation to the possible role that VIMFs, together with cortical MFs, may play in mediating cell surface focal contacts in the initial stages of cell attachment and spreading.
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  • 125
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    Cell Motility and the Cytoskeleton 13 (1989), S. 83-93 
    ISSN: 0886-1544
    Keywords: retinal pigment epithelium ; cytoskeleton ; focal contacts ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Retinal pigment epithelial (RPE) cells maintained in organ culture on Bruch's membrane and the associated choroid spread and migrate into a linear wound along the exposed basal lamina. Changes in cell shape, in the organization of microfilaments, and in cell-cell and cell-substratum interactions during this time were examined by epifluorescence and transmission electron microscopy. In contrast to cuboidal stationary cells distant from the wound edge, which display well-developed apical circumferential microfilament bundles (CMBs) associated with zonulae adhaerentes junctions, the migrating RPE cells near the wound edge instead are flat, and, in addition to microfilament bundles near junctions between adjacent cells, display prominent stress fibers. Furthermore, monoclonal antibodies to vinculin labeled regions at the terminal ends of these stress fibers indicating that the RPE cells form focal contacts with the basal lamina at these sites. Electron microscopy of these regions of cell-substratum interaction confirmed the presence of microfilament bundles that terminate on the cell membrane. Folds present in the basal lamina near these sites suggest that tension is being generated by the microfilaments in the stress fibers as the migrating cells pull on the underlying basal lamina through these adhesion points.
    Additional Material: 7 Ill.
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  • 126
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    Cell Motility and the Cytoskeleton 13 (1989), S. 181-194 
    ISSN: 0886-1544
    Keywords: desmosomes ; keratinocytes ; tumor cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In cultured human keratinocytes (NHEK) maintained in medium containing low levels of Ca2+ (0.04 mM) desmoplakin is a component of certain electron-dense bodies in the cytoplasm. These bodies are associated with bundles of intermediate filaments. Upon elevation of the level of Ca2+ in the culture medium to 1.2 mM, desmoplakin first appears at sites of cell - cell contact in association with bundles of intermediate filaments. Subsequently, desmoplakin becomes incorporated into desmosomes in a manner comparable to that seen in mouse keratinocytes (Jones and Goldman: Journal of Cell Biology 101:506-517, 1985). NHEK cells maintained for 24 hr at Ca2+ concentrations between 0.04 mM and 0.18 mM were processed for immunofluorescence, immunoelectron, and conventional electron microscopical analysis. In NHEK cells grown at Ca2+ concentrations of 0.11 mM, desmoplakin appears to be localized in electron-dense bodies associated with intermediate filaments at sites of cell - cell contact in the absence of formed desmosomes. At a Ca2+ concentration of 0.13 mM desmoplakin is arrayed like beads on a “string” of intermediate filaments at areas of cell - cell association. At 0.15 mM, desmosome formation occurs, and desmoplakin is associated with the desmosomal plaque. In basal cell carcinoma cells desmoplakin is not restricted to desmosomes but also occurs in certain electron-dense bodies morphologically similar to those seen in NHEK maintained in low levels of Ca2+ and during early stages of desmosome assembly. We discuss the possibility of “cycling” of desmoplakin through these bodies in proliferative cells.
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  • 127
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    Cell Motility and the Cytoskeleton 14 (1989), S. 156-162 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Additional Material: 2 Ill.
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  • 128
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 129
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    Cell Motility and the Cytoskeleton 14 (1989), S. 80-80 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 130
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989) 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 131
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989) 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 132
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 1-15 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: In order to develop a new kind of filter that makes a compromise between quasi-optical and more traditional bandpass filtering techniques at millimetre wavelengths, a theoretical modelling of grids in oversize and monomodal waveguides is performed using a variational method.
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  • 133
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 31-52 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: A novel procedure for the numerical modelling of current transport in semiconductor devices is presented. The method is based on high-order trigonometric expansions (Fourier series) of the solution. The expansion coefficients are calculated in a Galerkin-type algorithm. The method offers infinite-order accuracy regardless of the number of spatial dimensions of the model. Well-conditioning and diagonal dominance of the discrete equations render the numerical procedure stable and effective. Significant advantages are expected, particularly for the solution of strongly non-linear multidimensional device models. Properties of the algorithm are demonstrated on standard semiconductor devices.
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  • 134
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 227-240 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The coupling between incident electromagnetic fields and lossless, multimode, multiconductor, transmission systems is studied using transmission-line modelling (TLM). Model predictions compared well with experimental multimode results. The model presented is suitable for studying electromagnetic compatibility problems in the time-domain.
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  • 135
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 280-280 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 136
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 17-29 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The nature and significance of time-domain approximations needed for a diakoptic solution using transmission-line modelling (TLM) are examined. Truncation, filtering and repeated convolution of time responses result in severe restrictions in the accuracy of time-domain diakoptics. The frequency-domain diakoptic method has several advantages, including speed and simplicity, and warrants further study.
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  • 137
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989) 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 138
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 81-92 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: A de model for short-channel MOSFETs is presented in this paper. Several second-order effects associated with small-geometry MOSFETs such as mobility degradation, carrier velocity saturation and channel length modulation are included in the model. The analysis emphasizes the modeling of the output conductance and the transconductance which are important in analogue circuit simulation. The theoretical predictions of the model are in good agreement with the experimental data available in the literature.
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  • 139
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 75-80 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: A simulation of power transformers following switching operations is presented in which each core section is represented by a non-linear inductance, the differential inductance being determined from the flux density in that section of the core. In analysing the resultant circuit model for the transformer it is shown that the circuit equations have dyadic properties which can be used to give a more efficient numerical solution.
    Additional Material: 5 Ill.
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  • 140
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 93-102 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: An approach, based on the finite element method (FEM), is used for evaluating the electromagnetic field distribution in a class of E-plane metal insert filters, operating in the Ka-band. Artificial boundary conditions are inserted directly in a functional, enabling a solution of the scalar wave equation by finite element method.The scattering parameters of the filter, the electromagnetic field and, therefore, the current distributions on the thin metal inserts are computed. Then, the power dissipated in the metallic septa, which is one of the major limitations for the maximum CW-power that can be handled by these types of filter, is evaluated using a perturbational approach. Finally, the maximum field intensity is determined, allowing an estimate of the maximum peak power that these structures can handle.
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  • 141
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 103-111 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The multigrid technique, which is mainly used in the finite difference method, is applied to the TLM (transmission-line matrix or modeling) procedure. In the multigrid TLM, the field region is covered with sets of regular transmission-line matrices of different mesh sizes. The finer grid pattern, which overlaps the coarser grid, fills the region with high field gradients. Data obtained in the coarser grid are transferred to the finer grid through an interpolation process and used as boundary conditions for the latter. The finer grid data are then transferred back to the coarser one, thus improving the coarse grid values. The method is applied to several diffusion problems and significant improvements in accuracy and efficiency are shown.
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  • 142
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 115-115 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 143
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 131-142 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: Numerical quadratures encountered in solving integral equations and in finite element analysis often involve singular integrands, or integrands with very rapid local variation whose numerical stability resembles that of singularities. It is shown that specialized quadrature formulae of Gauss-Christoffel type can be generated for such integrands, using a recursive procedure based on the properties of the underlying orthogonal polynomials. Quadrature formulae of moderately high degree can be computed rapidly enough to allow them to be constructed as needed. An algorithm for generating quadrature formulae is given in detail. Singular or near -singular functions encountered in finite element analysis typically require three -point or four -point quadratures; such formulae are readily obtained, to seven -figure accuracy, in computing times short enough to regard the formulae as disposable and not worth preserving in tables.
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  • 144
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 187-188 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 145
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 189-190 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 146
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 191-201 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: This paper presents an overview of the 3D transmission-line matrix method of analysis, describing the various nodes which are available to date: expanded node, asymmetrical condensed node, symmetrical condensed node. The most general scattering matrices of these nodes are given. The various techniques used to implement irregularly graded meshes are also described. A comparison of all the processes applied to the same structure is made. Conclusions on their use are then discussed.
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 279-279 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 148
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 249-265 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The application of the transmission-line modelling technique to the design of semiconductor lasers and optical systems is reviewed. Generalized scattering matrices are developed to allow the technique to be applied to novel devices and systems.
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  • 149
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 61-73 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: A new P-N junction device model is presented. It consists of two numerical unidimensional macroscopic models. The first is a drift-diffusion model. The second is an energy model accounting for the carrier energy relaxation effects. Because device optimization requires systematic investigations, the computing time is of paramount importance and, so an accurate comparison between these two models was undertaken under both static steady-state and dynamic 94 GHz conditions in order to point out the relative interest of the energy model. This study is mainly devoted to silicon millimetre-wave IMPATTs. This comparison has highlighted the superiority of the energy model for the description of low field transport phenomena. But as IMPATT RF properties are mainly governed by high field transport, the RF power levels predicted by the two models have been found to be similar. Moreover it has shown that the energy model tends to underestimate both the efficiency and the diode terminal negative resistance level. This parasitic effect is inherent to the description of the carrier generation process by impact ionization which in this model is assumed to be governed by the carrier energy.
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  • 150
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 113-113 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 151
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 117-129 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The response of capacitive array sensors in the presence of flawed solid materials is simulated using finite elements and infinite elements with exponential decay. Conventional finite elements are used to model the critical regions near the probe and the surface of the solid. Infinite elements are used to represent the farfield conditions of the space surrounding the probe and the solid. The method is first applied to problems with analytic solutions to determine the accuracy of the results obtained using the infinite elements. The response of a capacitive array sensor is then simulated using a line integral which measures the relative change in admittance between flawed and unflawed solids. Examples of capacitive probe responses are given for several parametric variations of the flaw size and dielectric constant of the solid.
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  • 152
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 143-152 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The paper outlines the operating principles of a 3 -phase to single -phase cycloinverter, as used in heating and melting applications, and presents a digital computer model on which performance predictions may be based. Ideal power switches, an input filter and a compensated heating load are assumed in formulating an initial set of state -variable equations which are valid when all the switching devices are conducting. Connection matrix techniques are then used to generate the mesh equations relevant to the continually changing pattern of the switching devices, leading to a very versatile simulation program. Experimental results from a laboratory -scale unit are presented for different output power levels, and these are compared with corresponding computed results to illustrate the close similarity that exists.
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  • 153
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989) 
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    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
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  • 154
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 153-186 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: Rigorous full -wave analysis techniques are presently receiving much attention in the design of MICs and, in particular, of MMICs due to increasing circuit packing densities and structural complexity. In this paper, the problems associated with such techniques and previous related work are briefly outlined in the introduction. To obtain self -consistency, the electrodynamic Green's functions and related terms are then formulated for the shielded (M)MIC problem. The final operator equation for the numerical solutions derived and the associated functions space are presented and discussed. The central portion of the paper describes a new numerical solution using a discretized Green's function database technique. The resulting linear system of equations is solved by direct inversion for geometrical complexities involving up to about 1000 unknowns; for a higher number of unknowns, an iterative solution is generated. As an alternative to the database technique developed, a variety of spectral domain iterative solutions has been written and tested as well. This includes application of the conjugate gradient method to the normal operator equation (CGN algorithm), an implicit iterative Galerkin approximation called the modified planar conjugate gradient technique (MPCG) and monotonically convergent iteration procedure being a version of the conjugate residual algorithm (CR). Supplementary to this, the extraction of (M)MIC design data from the numerical 3D solutions obtained and error considerations are presented. The paper concludes with a variety of analysis examples of medium to high geometrical complexity and with verification of some results by comparison with measurements and with numerical data from other sources.CPU times required on typical workstations (Micro VAX, HP 9000, etc.) are moderate, thus rendering the techniques presented as useful in the solution of MIC and MMIC design problems.
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  • 155
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 53-60 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: This paper introduces a simple model for solving a system of first-order rate equations using the TLM numerical method. A general algorithm is developed by setting up a matrix of discrete compartments. The rate constants provide the linkage between each compartment and the neighbouring ones. The method is demonstrated by some examples in which saturation conditions are considered. The TLM solution is compared with analytical solutions and the Rung-Kutta method of solving differential equations.
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  • 156
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 203-214 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: This paper presents the dispersion characteristics of the FD-TD algorithm by showing the numerical phase and group velocities of the TEM, TE or TM modal solutions. For the TEM wave propagation, there exist three fundamental directions to which phase and group velocities can be expressed by only a single variable (wave number k or frequency ω). Those special directions were chosen to show the dispersive nature of the FD-TD algorithm of which the group and phase velocities depend explicitly on frequencies.In view of similarities between the TLM and FD-TD algorithms, a comparison of phase velocity characteristics of both methods was made. Under the special condition where the stability factor s is larger than 1/2, the FD-TD algorithm is found less dispersive than that of the original TLM. However, newly developed symmetrical TLM method appears less dispersive than the FD-TD algorithm.In the numerical simulation of waveguide modal solutions, it was found that there exists an optimum frequency which provide least numerical error in the FD-TD application. A procedure, which determines the stability factor s and the maximum allowed frequency Fmax for the best numerical results, is proposed in this paper.
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  • 157
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 241-248 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The TLM technique has been enhanced to allow the accurate modelling of EM diffusion through advanced composite material shells. This is a significant development owing to the rapidly increasing use of conducting composite materials in aerospace vehicle design. The accurate EM modelling of this class of materials is important for correct design of vehicles with regard to electromagnetic compatibility (EMC).Existing 3-D time-domain TLM models a non-perfectly conducting shell using a resistive sheet (or sheets) to simulate the surface. The results from this model are shown to be slightly inaccurate and require lengthy computer solution times. A separate 1-D, frequency-domain transmission-line model has been developed elsewhere to better analyse the diffusion properties of composite materials. The incorporation of the results from this frequency domain code into the 3-D time-domain TLM algorithm is outlined.
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  • 158
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 267-278 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: The calculation of the radar cross-section (RCS) of complex bodies using the symmetrical condensed TLM method is presented. The technique is based on a near-to-far field transformation of the TLM calculated near fields. Several two-dimensional examples are presented which validate the method. The main advantage of utilizing techniques such as TLM for RCS computation lies in the ability to model arbitrary bodies with complex material compositions.
    Additional Material: 11 Ill.
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  • 159
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    International Journal of Numerical Modelling: Electronic Networks, Devices and Fields 2 (1989), S. 215-225 
    ISSN: 0894-3370
    Keywords: Engineering ; Electrical and Electronics Engineering
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Electrical Engineering, Measurement and Control Technology
    Notes: A new TLM-based concept analogous to the Green's function approach in classical electromagnetic theory is presented. It employs the procedure known as time domain diakoptics. The response of a TLM mesh to a unit impulse excitation at selected input points is interpreted as a discrete Green's function in the time domain, and the term ‘Johns matrix’ is proposed for this characteristic response. As in classical theory, the response of the mesh to an arbitrary excitation is found by convolving the excitation with its Johns matrix. This concept extends the generalized scattering parameter concept into the time dimension, opens unprecedented possibilities for partioning time domain problems at the field level, and permits large-scale preprocessing of substructures for computer-aided design. It also represents and elegant way of modelling broadband absorbing boundary conditions, imperfectly conducting walls and general frequency dispersive boundaries in the time domain.
    Additional Material: 8 Ill.
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  • 160
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    Molecular Reproduction and Development 1 (1989), S. 98-106 
    ISSN: 1040-452X
    Keywords: Concatemers ; Ligation ; pRSVcat ; Recombination ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We describe preliminary experiments to analyse the fate of cloned DNA microinjected into the cytoplasm of the chick fertilised ovum. The reporter gene construct pRSVcat was injected into the germinal disc before the first cleavage divison, and the chick embryos were cultured for up to 7 days using the method of Perry (Nature 331:70-72, 1988). Linear plasmid molecules ligated rapidly after injection to form highmolecular-weight DNA molecules consisting mainly of random concatemers of the injected plasmid. Recombination involving circular molecules resulted in head-to-tail multimers of the plasmid. Some of the DNA was lost after injection, but the remainder was replicated approximately 20-fold during the first 24 h of development. Between days 1 and 7 in culture, the DNA was gradually lost and diluted out as the embryos developed. By day 7 in culture plasmid DNA was detectable in only 30% of the cultures analysed. No evidence for chromosomal integration of the exogenous DNA was obtained, suggesting that the plasmid DNA persisted episomally. Expression of the reporter gene construct pRSVcat was detected in day 2 and day 7 embryos.
    Additional Material: 6 Ill.
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  • 161
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    Molecular Reproduction and Development 1 (1989) 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 162
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    Molecular Reproduction and Development 1 (1989), S. 79-90 
    ISSN: 1040-452X
    Keywords: Cleaving embryo ; RNA synthesis ; DNA distribution ; Cattle ; Nucleogenesis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Eight-cell cow embryos were isolated and cultured in vitro in a medium enriched with 200 μCi of [5-3H]uridine for 20 min. Epon ultrathin sections of the embryos were investigated for the nucleolar morphology and for the appearance and localization of the sites of [5-3H]uridine incorporation by means of electron microscopic autoradiography. In addition to this, a general pattern of replicated embryonal DNA distribution was revealed by [methyl-3H]thymidine incorporation and light microscopic autoradiography.The essential phases of the transformation of the small nucleous precursor body (NPB) into a vast, functionally fully active nucleolus, characterized by typical nucleolar substructural components, are taking place within the eight-cell stage. This process differed in its morphology from the nucleologenetic process in early embryogenesis of other mammals, especially of that in the mouse.The first sign of NPB, transformation was the appearance of a large central vacuole followed later on by perinucleolar chromatin penetration into NPB, documented by both morphology and [3H]thymidine autoradiography. In some cases, concentration of dense fibrillar material forming clumps or stalks was seen in the central vacuole.The following rapid nucleolar development was characterized by the formation of secondary vacuoles concomitant with the onset of [5-3H]uridine incorporation into the dense fibrillar component and with the appearance of the first granules in the otherwise fibrillar structure of the nucleolus. During the late eight-cell stage, the still-rounded nucleolus developed features of a reticulated nucleolus known from somatic cells intensively synthesizing rRNA: a dense fibrillar component with associated labeling encircling fibrillar centers and a well-developed granular component. The labeled dense fibrillar component was observed mostly in the central area of the nucleolus; early embryonic NPB dense fibrous material not involved in transcription was disappearing rapidly. At the transition to the 16-cell stage the nucleoli lost their rounded shape because of the accumulation of a large amount of granular component, and they occupied a considerable part of the nucleus. In conclusion, the appearance of the nucleolar vacuole in eight-cell cow embryo is the starting point for following morphogenetic events linked with the onset of transcription.
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  • 163
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    Molecular Reproduction and Development 1 (1989), S. 138-145 
    ISSN: 1040-452X
    Keywords: Embryo ; In situ hybridisation ; Y probe ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: DNA probes are now widely used for prenatal diagnosis, but the prospect of preimplantation diagnosis of genetic disorders requires the development of sensitive genetic tests that can be performed on small numbers of cells removed from a preimplantation-stage pre-embryo. The sensitivity of molecular tests can now be increased by specifically amplifying the target DNA with the polymerase chain reaction. In situ hybridisation with chromosome-specific DNA probes to repeated sequences also permits the detection of particular numerical chromosome aberrations or the distinction of male and female pre-embryos when only a few interphase nuclei are available. We have used in situ hybridisation to a Y chromosome-specific DNA probe to sex preimplantation-stage pre-embryos and to sex fetuses from samples of chorionic villus cells, amniotic fluid cells, and fetal blood. These two approaches (amplification of target DNA and in situ hybridisation) provide suitable tests for improving prenatal diagnosis particularly when few cells are available and they offer the possibility of tests suitable for preimplantation diagnosis.
    Additional Material: 7 Ill.
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  • 164
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    Molecular Reproduction and Development 1 (1989), S. 249-253 
    ISSN: 1040-452X
    Keywords: Polyspermy ; Protein synthesis ; Human egg ; Fertilization ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: To investigate the early molecular events in human oocytes that are triggered by fertilization, the authors examined the pattern of polypeptides synthesized by unfertilized and dispermic embryos obtained through an in vitro fertilization and embryo transfer (IVF-ET) program. Compared with unfertilized oocytes of the same postovulatory age, the de novo protein synthesis in tripronuclear dispermic zygotes (21 hours postinsemination) was characterized by the appearance of three novel protein bands with molecular weights of 41.2, 35.3, and 26.0 kD. Concomitant with these changes, these zygotes showed the disappearance of bands at 54.0, 36.5, and 28.0 kD, along with the decreased synthesis of a protein band at 42.5 kD. Although 24% of the aged unfertilized oocytes exhibited bands corresponding to 41.2 and 35.3 kD, the 26.0 kD protein is restricted to the tripronuclear embryos. The significance of these results is discussed in relation to the use of polyspermic human oocytes as a model for the study of the early molecular events triggered by fertilization.
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  • 165
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    Molecular Reproduction and Development 1 (1989), S. 289-289 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 166
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    Molecular Reproduction and Development 1 (1989), S. 289-289 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 167
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    Molecular Reproduction and Development 1 (1989), S. 289-289 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 168
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    Molecular Reproduction and Development 1 (1989), S. 289-289 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 169
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    Molecular Reproduction and Development 1 (1989), S. 289-289 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 170
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    Molecular Reproduction and Development 1 (1989), S. 264-277 
    ISSN: 1040-452X
    Keywords: Nucleoproteins ; Element concentrations ; Electron microscopy ; Image analysis ; X-ray spectrophotometry ; Flow cytofluorometry ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: After cryosubstitution and Epon embedding, or after Nanoplast embedding and very thin sectioning, the chromatin of ejaculated or diluted boar spermatozoa appears to be formed of DNA fibers embedded in a quite homogeneous matrix. After sodium dodecyl sulfate (SDS) treatment, and to a lesser extent after freeze-thawing, the DNA fibers are present mostly between cords, probably proteinaceous in nature. The quantity of free sulfhydryl (SH) groups, as calculated from staining by DACM and flow fluorometry, is increased in thawed or SDS-treated cells. The quantity of NH2 groups, calculated from electron microscopy image analysis of alcoholic phosphotungstic acid-stained cells, is decreased in thawed nuclei. The DNA is more accessible to the fluorochrome ethidium bromide after freeze-thawing, and its sensitivity to HCI hydrolysis is modified, during the Feulgen-like staining procedure using acriflavine. The X-ray energy dispersive analysis of cryosections of nuclei indicates that the slight separation of DNA and nucleoproteins in freeze-thawed spermatozoa could result from a dramatic modification of the nuclear ionic environment during thawing.
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  • 171
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    Molecular Reproduction and Development 1 (1989), S. 283-288 
    ISSN: 1040-452X
    Keywords: Mouse/rat epididymis ; Acridine orange ; Disulfide bonding ; 7-Diethylamino-3-(4′-maleimidylphenyl)-4-methylcoumarin (CPM) ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Flow cytometric measurements were made on acridine orange (AO) and 7-diethylamino-3-(4′-maleimidylphenyl)-4-methylcoumarin (CPM)-stained epididymal- and vas deferens-derived spermatozoal nuclei to follow the course of chromatin condensation and oxidation of free sulfhydryl groups, respectively, during passage through mouse and rat posttesticular reproductive tracts. Alterations of mouse and rat spermatozoal chromatin during transition from a testicular elongated spermatids to epididymal caput spermatozoc resulted in a threefold loss of DNA stainability with AO. Passage of spermatozoa from the caput to corpus epididymis was accompanied by an approximate 15% loss of DNA stainability, which was maintained at that level throughout passage into the vas deferens. AO stainability of epididymal spermatozoal nuclei was generally independent of -SH group stainability. CPM stanability of rat spermatozoal nuclei free -SH groups was 83%, 18%, and 11% of caput spermatozoal values for corpus, cauda epididymis, and vas deferens, respectively. Comparable values for mice were 69%, 20%, and 18%. CPM stainability was relatively homogeneous for these mouse and rat reproductive tract regions, except mouse corpus epididymis spermatozoal nuclei stained very heterogeneously. Rat spermatozoa detained by ligature up to 7 days in the caput, corpus, and cauda epididymi had CPM staining values equal to or below those of normal vas spermatozoa, indicating that disulfide (S-S) bonding is intrinsic to the spermatozoa and is independent of the epididymal environment. These data suggest that chromatin condensation and loss of spermatozoal DNA stainability during passage from the testis to the vas deferens are independent of S-S bonding. Furthermore, the results are in agreement with previous findings suggesting that autoxidation of SH groups occurs independently of movement through the epididymis.
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  • 172
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    Molecular Reproduction and Development 1 (1989), S. 91-97 
    ISSN: 1040-452X
    Keywords: Protamines ; Disulfide bonds ; Osmium ammine reaction ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The arrangement of mouse sperm nuclei chromatin and, in particular, of DNA has been studied by electron microscopic cytochemistry. It had been previously shown that, after a Feulgen-type reaction using an osmium ammine complex (OAC), the OAC-stained DNA was distributed in a spotted pattern in the nucleus (Biggiogera: Basic Appl Histochem 30:501-504, 1986). The present chapter shows that this pattern is characteristic of mouse spermatozoa from testis to vas deferens, with the exception of some testicular spermatozoa, in which DNA was homogeneously stained. DNase digestion of thin-sectioned nuclei resulted in a distribution of residual material complementary to the pattern of the unstained zones after the OAC reaction. These findings are discussed considering the role of -S-S- crosslinks, characteristics of this extremely condensed chromatin, in limiting the availability of DNA to acid hydrolysis.
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  • 173
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    Molecular Reproduction and Development 1 (1989), S. 116-121 
    ISSN: 1040-452X
    Keywords: Bkm sequences ; Gonad differentiation ; Riboprobes ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The possible role of GATA/GACA repeated sequences in mammalian sex determination was investigated using Northern analyses of mouse and human RNA. Brain, liver, and gonadal RNA from three developmental stages of mice of both sexes and also human fetal RNA from various tissues were hybridized to both sense and antisense Bkm riboprobes as well as to the synthetic oligonucleotide (GATA)5. At low levels of stringency, putative transcripts of various sizes were observed in all tissue samples with all probes. At high stringency, only a putative transcript of approximately 12 kb was observed, but this was later shown to consist of contaminating DNA. No sex-specific differences were observed in any tissue or developmental stage. Thus, we find no evidence that the GATA/GACA repeated sequences are specifically expressed in quantities detectable by Northern analyses in a manner important to mammalian sex determination.
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  • 174
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    Molecular Reproduction and Development 1 (1989), S. 122-128 
    ISSN: 1040-452X
    Keywords: Pachytene spermatocytes ; Round spermatids ; Insulin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In vitro rat germ cell RNA synthesis is influenced by growth factors. Basic fibroblast growth factor (0.1 to 100 ng/ml) increases [3H]uridine incorporation in round spermatids (RS) but not in pachytene spermatocytes (PS); this effect is potentiated by insulin (10 μg/ml) and blocked in the presence of Sertoli cell-secreted proteins (SCSP). Somatomedin C (0.1 to 100 ng/ml) exhibits a similar effect when used alone without an influence by SCSP. Transforming growth factor β (0.1 to 10 ng/ml) acts on both cell types, but SCSP amplify this effect only in PS. These data suggest that growth factors synthesized in situ may play a role in the germ cell development and that their effects are moduiated by SCSP.
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    Molecular Reproduction and Development 1 (1989), S. 146-146 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 176
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    Molecular Reproduction and Development 1 (1989), S. 146-147 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 177
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 178
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    Molecular Reproduction and Development 1 (1989), S. 147-147 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 179
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    Molecular Reproduction and Development 1 (1989), S. 149-155 
    ISSN: 1040-452X
    Keywords: Gene regulation ; Echinoderm ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We report the first measurements of cell number, total RNA, and transcript accumulations for two actin genes during larval development of the sea urchin Strongylocentrotus purpuratus. At 5 weeks of feeding, when development of laboratory-raised larvae is completed, the cell number has increased about 100-fold with respect to the pluteus-stage embryo to about 150,000 ± 50,000, and the total RNA has increased 46-fold to about 130 ng per larva. The transcripts of the Cylla cytoskeletal actin gene, which is expressed in adult tissues, continue to accumulate throughout larval development. A contrasting pattern of transcript accumulation is observed for Cyllla, a different cytoskeletal actin gene that in the embryo is expressed only in aboral ectoderm. These transcripts increase in number early in larval development, when the larval epidermis is differentiating, and then decline in quantity. It is known that at metamorphosis the larval epidermis is largely histolyzed and that the Cyllla gene is not expressed in the juvenile or adult.
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  • 180
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    Molecular Reproduction and Development 1 (1989), S. 170-181 
    ISSN: 1040-452X
    Keywords: Multigene family ; Sequence analysis ; Developmental expression ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: To study the molecular aspects of the regulation of transcription of a multigene family, we have isolated and sequenced cDNA and genomic clones coding for the α-tubulin of the sea urchin Paracentrotus lividus. Two cDNA clones, Pα 10 and Pα 4, contain respectively the coding information for 391 C-terminal and for 338 N-terminal amino acids of the 452 residues that constitute the complete protein. They show silent nucleotide substitutions only, suggesting that Pα 10 and Pα 4 represent the cloned copies of two allelic gene transcripts, which encode for two α-tubulin isoforms with identical amino acid sequence in the region of the overlap. The comparison of the predicted amino acid sequence of the composite Pα 4-10 and of the mouse M α-6 (Villasante et al., Mol. Cell Biol 1986; 6:2409-2419) reveals a conservation of 97% between the two polypeptides. By RNA blotting hybridization six major α-tubulin transcripts were identified. Two, of 3.5 kb and 2.0 kb, are expressed in the unfertilized eggs and during early cleavage. The other two maternal mRNAs, of 2.4 kb and 1.8 kb, are expressed in both early and late cleavage embryos, but in the intestine the 1.8 kb RNA, which specifically reacted with the 3′ specific probe of the Pα 10 cDNA, is the only transcript detected. Finally, the 1.5 kb and 1.9 kb mRNAs represent the transcription of stage- and tissue-specific genes, respectively. In fact, the former becomes detectable at blastula stage and accumulates during late development, whereas the latter is found in the testis only. The sequence data of the 3′ terminus of the α-3 genomic clone suggests that it encodes for a divergent α-tubulin, and it most probably corresponds to the testis-specific gene.
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  • 181
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    Molecular Reproduction and Development 1 (1989), S. 201-207 
    ISSN: 1040-452X
    Keywords: Spermatozoa ; Human ; In situ hybridisation ; Y chromosome ; Sperm selection ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In situ hybridisation of a Y chromosome-specific DNA probe to preparations of decondensed spermatozoa revealed approximately 46.7% labelled spermatozoa among 3,900 scored. This is not significantly different from the 50% expected if only the Y chromosome-bearing spermatozoa are hybridised. Control hybridisations of Escherichia coli DNA and salmon testis DNA to decondensed sperm produced no significant labelling, whereas more than 99% of the spermatozoa were heavily labelled after hybridisation to total human DNA. These controls indicate that the methodology described in this paper renders the chromatin accessible for hybridisation and that the 50% hybridisation observed with the Y chromosome DNA probe was specific. In situ hybridisation with the Y probe therefore identifies the Y-bearing spermatozoa, and the protocol described should prove useful in evaluating methods of separating Y-bearing and X-bearing spermatozoa.
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  • 182
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    Molecular Reproduction and Development 1 (1989), S. 233-241 
    ISSN: 1040-452X
    Keywords: Preimplantation ; Gene expression ; RNA quantity ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Studies of gene expression during blastocyst formation in mouse preimplantation development have been limited by the amount of RNA available per embryo. Our present approach to this problem has been to construct a large, representative, blastocyst cDNA library in λgtll. Random hexadeoxynucleotides were used as primers with total blastocyst RNA serving as template. RNA collected from 4,100 32-64 cell embryos was used to generate a library with an initial size of 30 × 106 recombinants. By using clone frequency as a measure of relative mRNA abundance, our data support previous work on the relative and absolute amounts of actin, histone H2a, and intracisternal A particle. Furthermore, we provide estimates for the abundance of cytokeratin endo A, cytokeratin endo B, and β-tubulin from clone frequency data. Insert sizes for isolated clones range from 200 bp to 3.6 kb with full-length or near-full-length insert sizes for selected clones, indicating that random primer methods generate cDNAs which can represent a significant portion of the mRNA. We have so far characterized products whose abundance is equal to or greater than 0.002% of total RNA. This library offers the potential for the analyses of presumptive regulatory gene products in the mouse preimplantation embryo which are represented as low abundance (〈1% of mRNA) RNAs.
    Additional Material: 2 Ill.
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  • 183
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 156-163 
    ISSN: 1040-452X
    Keywords: Mitochondrial DNA ; Microinjection ; Embryogenesis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A microinjection procedure to introduce “paternal” mitochondria from a source other than spermatozoa into fertilized mouse eggs is described. When a mitochondrial suspension isolated from the testes or liver of Mus molossinus mice was microinjected into fertilized eggs of CD1 mice, the microinjected zygotes survived, developed normally, and offspring were produced. Mus molossinus mitochondrial DNA can be distinguished from CD1 mitochondrial DNA by Southern blot analyses using restriction enzymes such as Eco R1, Xba 1, or Spe 1. Although up to 120 viable mitochondria were injected, no exogenous mitochondrial DNA was detected in fetal samples or in the brain, liver, heart, testis, or ovary of the mature progeny. Under the experimental conditions used, similar results were obtained when mitochondria from the testes of New Zealand black mice or from testes of Syrian hamsters were microinjected into fertilized CD1 mouse eggs. Failure to detect the exogenous mitochondrial DNA under our assay conditions suggests that microinjected mitochondria from testis or liver did not selectively replicate during embryonic development. The “foreign” mitochondria appear to have the same fate during early embryogenesis as the mitochondria of the spermatozoon.
    Additional Material: 5 Ill.
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  • 184
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 182-192 
    ISSN: 1040-452X
    Keywords: Neurochordins ; Differentiation ; Embryogenesis ; Adenohypophysis ; Notochord ; Glycoconjugates ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: In a search for antigens immunologically related to chordin, a notochord-specific glycoprotein of sturgeneous fishes, extracts from 55 samples of human and rabbit tissues were tested for inhibition of [125I]chordin binding to rabbit polyclonal antibodies. The strongest inhibition was observed with brain extracts of both species. Human, chicken, rabbit, and newt brain extracts also inhibited chordin binding in liquid phase to monoclonal antibodies (MAbs) against the P-epitope, the most immunogenic epitope of this glycoprotein. Immunohistochemical studies done on human and chicken embryos, newt, sterlet, and sturgeon embryos, larvae, and juveniles revealed a strong immunoreactivity of the brain, spinal cord, and tissue of the peripheral nervous system with an anti-P MAb. Other tissues, with several exceptions, showed a negative reaction in immunohistochemical experiments. The authors found that the P-epitope is ontogenetically expressed in the neural tissue of chicken, newt, and sterlet at the period of cytodifferentiation. Gel chromatography of human, chicken, and newt brain extracts showed that in each case the P-epitope was associated with a polydisperse macromolecular material of similar size. These antigens were designated as neurochordins. Prolonged pronase digestion of human and chicken brain extracts resulted in fragments with M about 3 kDa (presumably glycopeptides), which reacted with anti-P MAbs. These fragments were of the same size as corresponding glycopeptides of the pronase digest of chordin. Thus, in the present study, the P-epitope has been shown to be characteristic for the neural tissue of several vertebrate species; in the brain, it has been found in association with neurochordins, macromolecular antigens that are presumably protein conjugates with carbohydrates.
    Additional Material: 7 Ill.
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  • 185
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 231-232 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 186
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 208-218 
    ISSN: 1040-452X
    Keywords: Embryonic regulation ; Nucleotide sequence ; Cylllb actin gene ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The developmental pattern of expression of the Strongylocentrotus purpuratus Cylllb actin gene was determined by RNA blot hybridizations carried out with a gene-specific probe and total embryonic RNA isolated from various stages of development. The results indicate that the Cylllb mRNA is not detected in the maternal pool, and, although the gene is activated at the early stages (about 10 hr postfertilization), considerable amounts of mRNA do not accumulate until well into the pluteus stage 3 days later. These results suggest either a post-transcriptional regulatory mechanism that governs early embryonic expression of the Cylllb actin or a late embryonic transcriptional enhancement of this gene. We present here the complete nucleotide sequence of the Cylllb gene, which lies within the 10,361 base pairs of the sequenced region. The entire transcription unit is 7,455 nt long and shares structural similarities with the other cytoskeletal-type actin genes from this sea urchi. Sequence comparisons of Cylllb to the Cyllla actin gene, to which it is linked, reveals extensive homology even in the introns. The deduced amino acid sequence of the Cylllb actin shows five amino acid substitutions compared with the Cyllla actin and nine when compared with the Cyl, the endodermal embryonic cytoskeletal-type actin. Five out of these nine amino acid differences occur within a small peptide (position 257 to 267). The 5′ flanking sequence of the Cylllb gene shows a remarkable homology (∼75-80%) with the Cyllla upstream region up to the position -200 and a lack of any obvious similarity further upstream. This observation suggests that the two genes possibly share some common regulatory factors.
    Additional Material: 5 Ill.
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  • 187
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 242-248 
    ISSN: 1040-452X
    Keywords: Transgenic mouse ; PCR ; Microinjection ; Bisection ; Morula ; Fetus ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: A polymerase chain reaction (PCR) system to detect transgenes in mouse preimplantation embryos was employed so that transgenic embryos could be selected before they were transferred to recipient mice. The selection system involves bisection of morulae, selection of the half-morulae containing target sequences within 7 hr, and culture and transfer of the sister half-morulae. PCR analysis of morulae derived from transgenic mice confirmed that the PCR system was reliable. However, five of 41 implanted embryos derived from PCR-positive morulae did not contain the transgenes. Also, one of 28 implanted embryos from PCR-negative morulae were transgenic. The selection system was applied to fertilized mouse eggs into which pSV2-gpt-gE1A DNA was injected. The injected DNA was detected in 30 of 84 morulae derived from the microinjected eggs. All seven implanted embryos developed from PCR-negative morulae had no detectable amount of transgenes, and one of two successfully implanted embryos from PCR-positive morulae was transgenic.
    Additional Material: 6 Ill.
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  • 188
    ISSN: 1040-452X
    Keywords: Retinoic acid treatment ; HOX gene clusters ; EC cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: We have studied the expression of four human homeobox genes representative of four different clusters (i.e., HOX-1, HOX-2, HOX-3 and HOX-5) in the embryonal carcinoma (EC) cell line NT2/D1. Following treatment with retinoic acid (RA), these cells differentiate into several cell types, including neurons, and steadily accumulate polyadenylated transcripts derived from the genes in a period ranging from 18 hr to 14 days of RA treatment. The sizes of major transcripts in differentiated EC cells coincide with those previously detected by the same probes in human embryos. Nuclear run-on transcriptional analysis showed no difference in the transcription rate of the four homeobox genes in differentiated vs. undifferentiated EC cells. Inhibition of protein synthesis by 5-18 hr of treatment of undifferentiated cells with cycloeximide causes accumulation of some homeobox transcripts at levels comparable to those observed after 18 hr of RA induction, although it does not cause superinduction in fully differentiated cells. These data suggest that the activation of homeobox gene expression in RA-induced EC cells is controlled, at least in part, by posttranscriptional mechanisms.
    Additional Material: 4 Ill.
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  • 189
    ISSN: 1040-452X
    Keywords: Y probe ; Prenatal diagnosis ; Mosaicism ; Chorionic villus biopsy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Samples of human adult lymphocytes, fetal lymphocytes, amniotic fluid cells, and chorionic villus cells were sexed independently by cytogenetics and DNA-DNA in situ hybridisation to a tritiated Y probe. For the in situ hybridisation analysis, the presence of Y bodies (hybridisation bodies) in 100 interphase nuclei were scored after autoradiography. In all, 82/83 samples were sexed in this way (one technical failure) and 78/82 were sexed by both in situ hybridisation and cytogenetics. There was complete agreement between the two methods. There was a considerable variation (40-100%) in the percentage of interphase nuclei with a hybridisation body among the male samples, but very few nuclei from female samples showed significant hybridisation. In situ hybridisation could be used to sex the conceptus when males but not females are at risk for various X-linked genetic disorders and may also be useful for detecting 45,X/46,XY mosaicism or polyploid/diploid mosaicism. This would be particularly useful for direct preparations of chorionic villus samples, which often prove difficult to analyse cytogenetically but offer the best means of avoiding maternal contamination. Some interphase nuclei had more than one hybridisation body, and this was most commonly found among amniotic fluid cells. Comparison of sizes of nuclei with one or two hybridisation bodies strongly suggested that most of the amniotic fluid cell nuclei with two hybridisation bodies were tetraploid.
    Additional Material: 6 Ill.
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  • 190
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 146-146 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 191
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 146-146 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 192
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 164-169 
    ISSN: 1040-452X
    Keywords: Growth Hormone-Releasing Factor ; Metallothionein-I ; Lambs ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Pronuclei of fertilized sheep ova were injected with fusion genes consisting of the mouse metallothionein-I promotor/regulator ligated to either the structural gene for bovine growth hormone (mMTbGH) or to a minigene for human growth hormone-releasing factor (mMThGRF). From a total of 842 sheep ova injected with mMTbGH and transferred into recipient ewes, 47 lambs were born. Two of the lambs were transgenic with mMTbGH, and both had bGH mRNA present in liver, kidney, and gut. In one lamb, plasma growth hormone was as high as 700 ng/ml. From a total of 435 sheep ova injected with mMThGRF and transferred to recipients, 54 lambs were born and 9 fetuses were collected. Nine of the 63 had integrated the mMThGRF gene. One of the nine had high concentrations of immunoassayable hGRF in its plasma and high variable plasma concentrations of ovine growth hormone. The lamb that expressed the hGRF gene did not release GH in response to an hGRF challenge. Four of five fetal offspring of a nonexpressing mMThGRF transgenic ram also contained the mMThGRF gene and, like the sire, failed to express the gene as determined by either liver hGRF mRNA or by plasma hGRF. Growth of the single transgenic lamb expressing hGRF was similar to control lambs. These studies demonstrate efficient introduction of genes into the sheep genome and indicate that transgenes are expressed and heritable.
    Additional Material: 4 Ill.
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  • 193
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 193-200 
    ISSN: 1040-452X
    Keywords: Gene expression ; Testis ; Protamine ; DNA binding proteins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Transition protein 1 (TP1) is a small basic nuclear protein that functions in chromatin condensation during spermatogenesis in mammals. Here, recently identified cDNA clones encoding mouse transition protein 1(mTP1) were used to characterize the expression of the mTP1 mRNA during spermatogenesis. Southern blot analysis demonstrates that there is a single copy of the gene for transition protein 1 in the mouse genome. Northern blot analysis demonstrates that mTP1 mRNA is a polyadenylated mRNA approximately 600 bases long, which is first detected at the round spermatid stage of spermatogenesis. mTP1 mRNA is not detectable in poly(A)+ RNAs isolated from mouse brain, kidney, liver, or thigh muscle. mTP1 mRNA is translationally regulated in that it is first detected in round spermatids, but no protein product is detectable until approximately 3 days later in elongating spermatids. In total cellular RNA isolated from stages in which mTP1 is synthesized, the mTP1 mRNA is present as a heterogeneous class of mRNAs that vary in size from about 480 to 600 bases. The shortened, heterogeneous mTP1 mRNAs are found in the polysome region of sucrose gradients, while the longer, more homogeneous mTP1 mRNAs are present in the postmonosomal fractions.
    Additional Material: 5 Ill.
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  • 194
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989) 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 195
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 219-229 
    ISSN: 1040-452X
    Keywords: Testis ; Nutritive phagocytes ; Male germ line cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The expression of testis-specific and adult somatic histone genes in sea urchin testis was investigated by in situ hybridization. The testis-specific histone genes (Sp H2B-1 of Strongylocentrotus purpuratus and Sp H2B-2 of Lytechinus pictus) were expressed exclusively in a subset of male germ line cells. These cells are morphologically identical to replicating cells pulse-labeled with 3H-thymidine. Genes coding for histones expressed in adult somatic and late embryo cells (H2A-β for S. purpuratus and H3-1 for L. Pictus) were expressed in the same germ line cells, as well as in the supportive cells (nutritive phagocytes) of the gonad. All histone mRNAs detected in the male germ lineage declined precipitously by the early spermatid stage, before cytoplasmic reduction. The data suggest that both testis-specific and adult somatic histone genes are expressed in proliferating male germ line cells. Testis-specific gene expression is restricted to spermatogonia and premeiotic spermatid, but somatic histone expression is not. The decline of histone mRNA in nondividing spermatids is not merely a consequence of cytoplasmic shedding, but probably reflects mRNA turnover.
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  • 196
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 146-146 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 197
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 146-146 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
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  • 198
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989) 
    ISSN: 1040-452X
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Type of Medium: Electronic Resource
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  • 199
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 254-263 
    ISSN: 1040-452X
    Keywords: mRNA localization ; In situ hybridization ; Blastocysts ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The distribution of total polyadenylated RNA and mRNAs from the β-actin, fibronectin, and cytokeratin Endo A genes was examined in preimplantation mouse embryos using in situ hybridization of riboprobes to RNA in sections of embryos. Polyadenylated RNA was found in the cytoplasm of all cells of blastocyst-stage embryos, whereas the specific mRNAs displayed three distinct patterns of expression: uniform throughout the embryo (β-actin), enriched in the inner cell mass (fibronectin), and enriched in the trophectoderm (Endo A). In eight-cell embryos, the polyadenylated RNA was more concentrated in nuclei than in the cytoplasm (as noted previously), although this was not the case in blastocysts, nor was it true for the specific mRNAs that were examined. These experiments demonstrate that there is localized gene expression in the early mouse embryo, which correlates with the formation of the trophectoderm and the inner cell mass.
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  • 200
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    New York, NY [u.a.] : Wiley-Blackwell
    Molecular Reproduction and Development 1 (1989), S. 278-282 
    ISSN: 1040-452X
    Keywords: Vitellogenin ; Lipophorin ; Vitellogenic mRNA ; Heat shock ; Fat body ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Vitellogenin (Vg) and lipophorin (Lp) are synthesized by the fat body of adult locust (Locusta migratoria) females. We have shown by an immunohistochemical technique that both proteins are produced in the same cells of the fat body. The rate of Vg synthesis was measured with the use of double immunoprecipitation of labeled proteins at oviposition and 24 h later. It was found that the rate of Vg synthesis declined significantly by the time of oviposition; however, 24 h later, it was raised to the highest possible level. The rate of Lp synthesis remained constant at both indicated points. The similar postlaying increase in the Vg synthesis rate was observed in the fat bodies of females treated by α-amanitin immediately after oviposition. The data provide evidence that Vg biosynthesis in L. migratoria is regulated by selective periodical repression and derepression of Vg mRNAs in the fat body cells but not by total inhibition and stimulation of protein-synthesizing machinery.
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