摘要
将烟草黄矮双生病毒（Ｔｏｂａｃｏｙｅｌｏｗｄｗａｒｆｇｅｍｉｎｉｖｉｒｕｓ，简称ＴｏｂＹＤＶ）中双向启动子的区域，以不同长度的片段和方向插入启动子分析载体ｐＧ１，与ＧＵＳ报道基因和ＮＯＳ终止子融合。同时，将各个ＴｏｂＹＤＶ读码框区域插入表达载体ｐＡＲＴ７中，置于ＣａＭＶ３５Ｓ启动子和ＯＣＳ终止子之间。用电穿孔法将各种启动子构建物个别地或者与读码框构建物成对地导入烟草和玉米原生质体，以考察ＴｏｂＹＤＶ启动子控制下ＧＵＳ基因瞬间表达的活性，以及ＴｏｂＹＤＶ的读码框编码产物对启动子活性的调控作用。结果表明，在烟草原生质体中，Ｃ１＋２基因的启动子活性最强，并受到Ｃ１＋２读码框编码产物的抑制；而Ｖ１和Ｖ２读码框的启动子活性较弱，然而它为Ｃ１＋２和Ｖ２，Ｖ２读码框构建物编码产物增强。在玉米原生质体中，与Ｖ１和Ｖ２基因的启动子相比。
DNA fragments from the bidirectional promoter region of the Tobacco yellow dwarf geminivirus (TobYDV) were cloned into pUC18-based vector pG1, producing transcriptional fusion with the β-glucuronidase (GUS) gene and nopaline synthetase terminator sequence. The activity for each promoter construct was analyzed by a GUS expression assay from N. clevelandii, N. plumbagfolia and Zea mays (maize) Black Mexican Sweet protoplasts extract or from N. clevelandii protoplast extract coelectroporated with the GUS reporter constructs in which TobYDV open reading frames (ORFs) replaced GUS gene individually and was controlled by a califlower mosaic virus 35S promoter. From tobacco protoplasts, strong promoter activity was observed for the promoter of C1+C2 gene which was repressed by coelectroporation of C1+2 ORF construct. Moderate activity was observed for the promoter of the V1 and V2 ORFs which were enhanced by coelectroporation of the C1+2, V1 and V2 ORF constructs. Whereas the activity of C1+2 gene promoter was weaker comparing to the activity of V1 and V2 gene promoters in maize protoplasts.
出处
《病毒学报》
CAS
CSCD
北大核心
1998年第1期68-74,共7页
Chinese Journal of Virology
基金
中国与新西兰学者交流基金
关键词
烟草
黄矮双生病毒
双向启动子
调节控制
Tobacco yellow dwarf geminivirus, Bidirectional promoter, Transient expression and regulation
