摘要
用冻融法将含有绿色荧光蛋白基因的质粒pGL2-GS导入农杆菌菌株LBA4404 (pTOK233),两个质粒发生同源重组,形成一个新的二元载体pTOK233-GS.农杆菌菌株LBA4404 (pTOK233-GS) 经叶盘法转化烟草,筛选具有卡那霉素抗性的愈伤组织,诱导成苗.PCR 扩增发现,绿色荧光蛋白基因在90% 的再生植株中存在.在荧光显微镜下,使用蓝色激发光观察再生植株的徒手切片和压片发现,80 % 以上的再生植株都能不同程度地发出绿色荧光.多数植株的根尖和叶脉都发光,一些植株的气孔。
Plasmid pGL2 GS containing gfp was constructed and introduced into A. tumenfaciens strain LBA4404(pTOK233). A new strain LBA4404(pTOK233 GS) was generated by homologous recombination between plasmids pTOK233 and pGL2 GS. Discs of tobacco leaves were transformed with LBA4404(pTOK233 GS) strain. Calli with kanamycin resistance and GUS activity were selected and induced to regenerate tobacco plants. PCR amplification showed that 90% of the Kan R plants contain gfp. The observation of gfp expression in tobacco plants with slides was carried out under fluorescent microscope upon excitation of blue light. It was found that more than 80% of plants shone green fluorescence of various strengthes. Most of them shone green fluorescence at bundle and root tips. In a few plants, green fluorescence could also be seen at air pore, epiderm or tomentum.
出处
《华中理工大学学报》
CSCD
北大核心
1999年第9期64-66,共3页
Journal of Huazhong University of Science and Technology
基金
国家自然科学基金
湖北省生物化学和分子生物学重点学科基金
关键词
基因表达
绿色荧光蛋白
烟草植株
农杆菌
gene expression
green fluorescent protein
tobacco plant
agrobacterium tumenfaciens Luo Zhongxun Prof.
College of Life Sci., Hubei University, Wuhan 430062, China.