【摘要】 结合Cre/loxp定位重组系统和杂种一代的育种特点,构建了雄性不育基因表达载体pCABARTABn和相应的恢复基因表达载体pBINPLUSCre。将位于质粒pTTABn中的雄性不育基因表达盒(含TA29启动子、barnase基因及Nos终止子)切下插入中间克隆载体pGloxp2个同向lox位点之间,再将上述表达盒连同lox位点切下插入pCAMBar,获得以除草剂Basta为筛选剂的雄性不育基因表达载体pCABARTABn。PCR方法从溶原菌BM25.8基因组DNA扩增出Cre基因,克隆测序验证无误后插入PBI525CaMV35S-35S双启动子和Nos终止子之间,再将表达盒切下插入pBINPLUS质粒相应位点,得到恢复基因植物表达载体pBINPLUSCre。更多还原

【Abstract】 The male sterility gene expression vector pCABARTABn and the fertility- restoring gene expression vector pBINPLUSCre were constructed by combining the features of the Cre/loxp site-specific recombination system and F1 hybrid breeding. The sterile gene expression box in pTTABn vector, which contained the TA 29 promotor, barnase gene and nos terminator, was cut down and then inserted in between the 2 directly oriented loxp sites of pGloxp. The final expression vector pCABARTABn was obtained by inserting the sterile gene expression box with the two directly oriented loxp sites into the plasmid pCAMBar, which contained the Bar gene. The Cre gene was cloned from lysogen BM 25.8 with the PCR procedure. The Cre gene was then inserted in between the CaMV 35 S-35 S double promoter and nos terminator of PBI 525 plasmid after sequencing. The expression box was cut down and inserted into plasmid pBINPLUS to obtain the final fertility-restoring gene expression vector pBINPLUSCre.更多还原