摘要
为了分离水稻的基因及其启动子,该实验室构建了T-DNA(GUS)结构的水稻启动子捕获系统,对其中编号为113#、T-DNA单拷贝插入、GUS报告基因为组成型表达的阳性捕获系进行了进一步分析。潮霉素筛选结合GUS组织化学染色获得了T-DNA插入的纯合株(113#-22和113#-26);Inverse法分离得到T-DNA插入位点水稻基因组DNA旁邻序列,测序和BLAST结果表明,T-DNA反方向插在水稻基因组4号染色体预测基因的内含子中;扩增T-DNA插入位点上游2kb左右DNA片段,构建启动子分析质粒转化水稻‘中花11’胚性愈伤组织,获得转基因植株,GUS组织化学染色模式与113#阳性株系一致。结果表明,该预测基因及其启动子是利用启动子捕获系统所捕获到的候选基因。
In order to isolate genes and their promoters in rice,we created a promoter trap system containing the promoterless β-glucuronidase （GUS） reporter gene in the T-DNA region.113# was further analysisied,in which the GUS reporter gene was expressed constitutively.A single copy of the T-DNA was inserted into the plant genome,and the flanking sequence around T-DNA was isolated by inverse PCR.Sequencing and BLAST analyses suggested the T-DNA was inserted reversely in a candidate gene of rice chromosome 4.The promoter of the candidate gene directed GUS expression constitutively,analogous to the GUS expression pattern observed in 113#.The results confirmed that the candidate gene was trapped by T-DNA （GUS） structure.
出处
《西北植物学报》
CAS
CSCD
北大核心
2013年第7期1298-1303,共6页
Acta Botanica Boreali-Occidentalia Sinica
基金
植物分子遗传国家重点实验室开放课题
江苏师范大学科研基金项目(10XLA06)
