ISSN:
1432-203X
Keywords:
Key words Gentian
;
Hygromycin phosphotransferase gene (hpt)
;
Particle bombardment
;
Transformation
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Abstract Cell suspension cultures were established from leaf explants of gentian (Gentiana triflora×G. scabra) for the generation of transgenic plants by particle bombardment. The parameters for the bombardment of suspension culture cells with a particle gun were examined by monitoring the transient expression of a gene for β-glucuronidase driven by the cauliflower mosaic virus (CaMV) 35S promoter. We found that prior culture of suspension culture cells for 5 days on solid medium was optimum for successful particle bombardment. Putative transformed calli were obtained from bombarded cells after a two-step selection procedure. Cells were cultured first with 30 mg l–1 hygromycin in liquid MS medium that contained 10 mg l–1 N-phenyl-N′-1,2,3-thiadiazol-5-yl urea, 1 mg/l 1-naphthaleneacetic acid and 30 g l–1 sucrose and then on solid medium prepared from the same liquid medium plus 2 g l–1 gellan gum. After 12 weeks of selection on solid medium that contained 30 mg l–1 hygromycin, two transgenic gentian plants were regenerated from each selected callus. Analysis by the polymerase chain reaction and Southern blotting revealed the stable integration of transferred DNA.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/s002990050755
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