摘要
[目的]建立大白菜的硝酸还原酶基因(NR)转化体系。[方法]以大白菜北京小杂60为材料,通过农杆菌LBA4404介导,将硝酸还原酶基因(NR)转入大白菜,对获得的转NR基因苗进行PCR检测、Southern杂交检测以及硝酸盐含量测定。[结果]试验初步建立了一套大白菜的转化体系,即以预培养2d苗龄4d的大白菜不带柄半子叶为外植体,在OD600nm值约为0.6的农杆菌工程菌液中浸染8min,然后在pH值为5.8且不加抗生素的分化培养基中将子叶与菌共培养2d,其后经过诱导、抗生素筛选、生根生长等过程,获得抗性植株,转化频率为6.48%。对抗性植株采用分级筛选,获得8株转NR基因苗。[结论]试验证实,NR基因已整合进大白菜的基因组中并得到表达,植株体内的硝酸盐含量均低于对照。
[ Objective ] The purpose was to establish the nitrate reductase gene （NR） transformation system for Chinese cabbage. [ Method ] With Chinese cabbage of Beijing Xiaoza 60 as material, the NR was transferred into Chinese cabbage through Agrobactcrium LBA4404 mediating, the transgenic seedlings obtained were conducted for PCR detection and Southern blot as well as nitrate content determination. [ Result] A transformation system of Chinese cabbage was founded preliminarily in the test ： with half cotyledon, without petiole of Chinese cabbages with 4 d seedling age, which were pre-cuhured for 2 d, as explants, they were infected in Agrobacterium culture medium with OD600 nm value about 0. 6 for 8 min, and then the cotyledon and Agmbacterium were co-cultured for 2 d in the differentiation medium without antibiotics with pH value of 5.8, afterwards, through the inducement, antibiotics screening, rooting and growth etc. , the resistant plants were acquired with the transformation frequency of 6.48%. The screen by grade was done on resistant plants and 8 transgenie NR seedlings were gained. [ Conclusion] The test validated that NR gene was integrated into the Chinese cabbage gene group and was expressed, and nitrate contents in plants were all lower than that of control ones.
出处
《安徽农业科学》
CAS
北大核心
2009年第30期14597-14599,共3页
Journal of Anhui Agricultural Sciences
基金
甘肃省科学事业费项目(QS022-C31-048)
