FACTORS INFLUENCIMG GENETIC TRANSFORMATION OF HYPOCOTYL PROTOPLASTS OF PHASEOLUS AUREUS (MUNG BEAN)
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摘要: 采用共培养法和PEG法将外源基因导入绿豆下胚轴原生质体中,得到转化愈伤组织。根癌农杆菌经乙酰丁香酮等诱导物质以及低pH、低PO43-预培养后,细菌T-DNA的转移受到促进,GUS基因表达活性明显提高。在PEG介导的绿豆原生质体转化中,高Ca2+/pH有利于外源DNA转移到植物细胞,GUS活性较高。此外,质粒DNA浓度和启动子类型均对外源基因的转移和表达有一定影响。Abstract: In this paper,we studied on factors influencing transformation of hypocotyl protoplasts of mung bean (Phaseolus aureus).The experimental result showed th t lower pH,phosphate starvation and acetosyringone could activate the expression of the vir genes of Ti-plasmid in Agrobacterium tumefaciens and promote T-DNA transfer into mung bean cells to some extent through transient expression of GUS activity in mung bean cells. Higher pH and concentration of Ca2+ promoted transient expression of GUS gene in PEG-mediated DNA direct transfer.The activity of GUS gene increased with the increase of foreign plasmid DNA concentration in the range of 10-100μg. The transformed calli from mung bean hypocotyl protoplasts were selected on MSO medium containing 50 mg/L chloramphanicol after the protoplasts were cocultivated with A.tumefaciens strain C58C1 (pBZ6111 ),and confirmed by analysis of nopaline synthetase (NOS) activity.
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Keywords:
- Phaseolus aureus /
- Protoplast culture /
- Transformation /
- Transient expression
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