rd29A启动子的克隆及提高烟草抗逆性的研究

Study on Cloning of rd29A Promoter and Enhancing Stress Tolerance of Tobacco

根据GenBank上公布的rd29A基因序列(D13044),利用PCR方法从拟南芥(Arabidopsis thaliana)基因组DNA中扩增得到了rd29A基因的启动子片段。序列分析表明,该片段与D13044有99%的同源性,它包括了DRE等4种完整的顺式作用元件。与GUS基因融合构建双元植物表达载体pBI-rd。转基因烟草的GUS活性组织染色分析及Northern杂交分析均表明,3种胁迫处理均可诱导GUS基因大量表达。其中,15%PEG和0.5%NaCl胁迫处理的转基因烟草比0℃低温处理的转基因烟草的GUS表达量高,而未经胁迫处理的转基因烟草的GUS基因只有少量表达。这些结果表明,rd29A属胁迫诱导型启动子,当植物遭受逆境胁迫时,rd29A启动子可以驱动下游目的基因超量表达,这就为通过基因工程途径提高植物抗逆性奠定了基础。

Promoter of rd29A from Arabidopsis thaliana was obtained by PCR with the primers designed according to the sequence of rd29A （D13044） which was published on GenBank. Sequence analysis indicated that this sequence was 99% identified with D13044 and there were four integrated cis-acting elements including DRE. Plant binary expression vector pBI-rd was constructed by fusing this rd29A sequence with GUS gene. Both analysis of GUS active tissue dying and Northern blot of transgenic tobacco showed that three stress treatments could induce GUS to express abundantly. 15% PEG and 0.5% NaC1 could induce GUS to express stronger than low temperature, while GUS expressed little in untreated transgenic tobacco. These results indicated that the promoter of rd29A was an inducible promoter. When plant was in various stresses, the promoter of rd29A could drive downstream target genes to be highly expressed. These findings can be further used in improving plant tolerance by gene engineering.