【摘要】 利用网络数据库http://jura.wi.mit.edu/bioc/siRNAext/,确定玉米淀粉分支酶SBEⅡb的部分基因序列作为RNAi的目标序列,通过RT-PCR方法从玉米自交系‘综31’中分别成功克隆出了玉米淀粉分支酶sbeⅡb基因的部分序列及其启动子序列,从大肠杆菌中成功克隆了木糖异构酶基因xylA作为筛选标记,构建了含有35S驱动的木糖异构酶基因和sbeⅡb启动子驱动的含有sbeⅡb目标序列的反向重复序列的表达载体pBAC413,并对玉米自交系实施转化。经PCR检测转基因玉米再生植株,初步判定目的序列已经成功整合到玉米再生植株的基因组中。更多还原

【Abstract】 As the targeted sequences of RNAi,parts of sbeⅡb genes encoding starch branching enzyme in maize was ascertained by searching internet database http:$$$$jura.wi.mit.edu/bioc/siRNAext/.Segments of sbeⅡb and their promoters existed in maize inbred line ‘Zong 31’ were cloned separately by RT-PCR.An expression vector of RNAi with the fragment of sbeⅡb sense gene+FAD2 intron+anti-sense gene was constructed,named as pBAC413,to regulate the biosynthesis of maize amylose.The xylA gene from Escherichia coli encoding xylose isomerase which is used as selective marker was cloned by PCR.Expression vector pBAC413 containing CaMV35S promotor driving xylA gene and reverse repeat sequence of starch branching enzyme gene (sbeⅡb) drived by its corresponding promotor was successfully constructed.pBAC413 was transformed with maize calli induced from the elite inbred lines.Successful integration of xylA gene into maize genome was confirmed by PCR.更多还原