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  • 1
    ISSN: 1432-0983
    Keywords: Aspergillus niger ; Adenine mutants ; Pyrimidine mutants ; Linkage groups ; Genetic analysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Mutants of Aspergillus niger requiring adenine and one mutant requiring cytosine were isolated after low-dose mutagenesis and enrichment. In addition we had mutants of two genes involved in the pyrimidine biosynthesis isolated as 5-fluoro-orotic acid-resistant mutants. The fifteen adenine-less mutants could be placed in seven complementation groups. From each group a representative mutant was analyzed in order to determine the linkage group by analysis of the mutants in a heterozygous diploid carrying markers in six linkage groups. AdeF could not be assigned to any one of these linkage groups and proved to be linked to nicB, oliC and cnxC, none of which could be placed in a linkage group. Thus, conclusive evidence was obtained for a seventh linkage group. As pyrA was used as selection marker for transformation, we constructed a pyrA strain with a linked marker which can be used in the genetic analysis of transformants.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-0983
    Keywords: Aspergillus niger ; Genetic analysis ; Genetic markers ; Linkage groups ; Master strains
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A start has been made on establishing a collection of Aspergillus niger colour and auxotrophic mutants with an isogenic background for use as a source of genetic markers. All strains have short conidiophores (cspAl ), which makes them easy to handle on test plates. Genetic markers were combined stepwise by somatic recombination. Somatic diploids were obtained at frequencies of 10−6-10−5 with conidiospores collected from a heterokaryon. The haploidization of heterozygous diploids was induced by benomyl. For unlinked markers, the frequency of recombinants varied from 35%–65%. Low frequencies of recombinants were found between markers on a same chromosome, but this was sometimes disturbed by mitotic crossing-over during an early stage of the diploid. Master strains were constructed having markers for six linkage groups.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-0983
    Keywords: Benzoate conversion ; Mutant isolation ; Transformants ; Aspergillus niger
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary This study was prompted by the observation that an Aspergillus niger transformant with a multicopy bphA (benzoate-4-hydroxylase gene) insert did not grow on benzoate, whereas a transformant with only one extra copy could grow. Therefore, an extensive survey has been made for other genes involved in the conversion of benzoate into 4-hydroxy-benzoate. A transformant with two copies of the bphA gene was used in part of the mutation experiments in order to avoid the isolation of many bphA mutants. Filtration enrichment was used to isolate mutants defective in the conversion of benzoate. The Bph mutants that have been isolated belong to six complementation groups. Mutants with a defected structural gene (bphA) were again predominantly found but, in addition, five other groups of mutants that could not grow on benzoate were isolated. Genetic analysis of the mutants showed that the six genes were localized in different parts of the genome. This was used as an additional proof that some mutants involved different genes. Diploids with seven copies of the bphA gene and heterozygous for one of the other bph genes were constructed. No indication has been obtained that any one of the mutant classes is responsible for the growth-limiting factor in bphA multicopy transformants. This study shows that the p-hydroxylation of benzoate is very complex, although the metabolic pathway is straight forward.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-0983
    Keywords: Aspergillus niger ; Auxotrophic mutants ; Filtration enrichment ; Novozym enrichment ; Lytic enzymes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary More than 100 auxotrophic mutants of Aspergillus niger were isolated using filtration enrichment. The mutants obtained in this way were predominantly aminoacid requiring. Hardly any vitamin-deficient mutants were found. The limitations of the method turned out to be due to cross feeding in liquid medium and non-specific loss of ungerminated conidiospores. To circumvent these problems an enrichment method has been used in which conidiospores that germinated on solid medium were selectively killed by the lytic enzyme preparation Novozym 234. We found auxotrophic mutants at high frequencies and several new types of mutants. Optimal conditions for the enrichment procedures have been determined. Essential factors appeared to be segregation of the conidia after mutagenic treatment, in order to obtain synchronization of germination, and the incubation time of the conidia on minimal medium prior to enzymic treatment. Under appropriate conditions Novozym enrichment proved to be very efficient. Depending on the type of mutants desired, one or both procedures can provide an effective method for the enrichment of mutants with a metabolic defect. The Novozym method can be adapted to other fungi and some of the observations that are described may also be of importance to improve other enrichment methods that are based on the selective killing of germinating conidiospores.
    Type of Medium: Electronic Resource
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