摘要
采用反转录－聚合酶链式反应方法（ＲＴＰ－ＣＲ），在人工合成的引物引导下，扩增出水稻矮缩病毒基因组第一片段（Ｓ１）全长序列及第五片段的部分序列（ＳＳⅢ）．将扩增的片段分别克隆到克隆载体ｐＧＥＭ７Ｚｆ（＋）及ｐＵＣ１９的ｓｍａｌ位点上，并进行了序列测定。在此基础上，利用ｐＣＲ引入的方法将核酶序列引入到Ｓ５Ⅲ片段反义链上以构成反义核酶基因Ｓ５ⅢＲ，将Ｓ１片段５＇端部分序列（Ｓ１－１）及Ｓ５ⅢＲ基因克隆到植物表达载体ｐＲＯＫⅡ上，构建成水稻转化载体ｐＲＯＫ－Ｓ１－１＇及ｐＲＯＫ－Ｓ５ⅢＲ。
he whole sequence of RDV S1 fragment and partial sequence(S5Ⅲ)of RDV S5 fragmentwere synthesized and amplified by the method of reversed transcription-polymerase chain reaction(RT-PCR).Each amplified product was then cloned into pGEM7Zf(+)or pUC19 at the Smalsite,and sequenced.Based on these results,a hammer-head ribozyme sequence was introduced intoantisense chain of S5Ⅲfragment to construct antisense ribozyme gene S5ⅢR,Rice transformationvector pROK-S1-1' and pROK-S5ⅢRwere constructed by inserting S5ⅢRgene and 5'-end se-quence of S1 fragment(S1-1')into plant expression vector pROK Ⅱ.
出处
《中国病毒学》
CSCD
1996年第2期164-169,共6页
Virologica Sinica
