摘要
从成熟的薄皮甜瓜(Cucumis melo cv.Qitian I)果肉组织中分离总RNA,经RT-PCR扩增得到0.7 kb的cDNA片段,将其克隆到质粒载体pGEM-T Easy。测序表明,该基因为777 bp,编码258个氨基酸,与Gen-Bank中甜瓜1-氨基环丙烷-1-羧酸合成酶(1-aminocyclopropane-1-carboxylate synthase,ACS)基因比对,核苷酸同源性为99%,氨基酸同源性为98%。克隆片段经双酶切消化,反向插入到植物表达载体pCAM2301的E8启动子和NOS终止子之间,构建成E8调控下ACS基因反义表达载体。通过冻融法将携带反义cDNA的植物表达载体质粒转入根癌农杆菌LBA4404,得到了完整的Ti质粒表达载体系统。
A cDNA fragment of an amino-cyclopropane-l-carboxylie acid （ACC） synthase was amplified by Reverse Transcription Polymerase Chain Reaction （RT-PCR） from sarcocarp tissue of mature nmskmelon （Cucumis melo cv. Qitian I） and cloned into a vector pGEM-T Easy. DNA sequencing showed that the cloned fragment shared highly homogeneity to the cDNAs of ACC synthases from other Cucubitae plants. The cloned eDNA of ACC synthase was further introduced into binary vector pCAM2301 in a reverse orientation with its upstream promoter （E8）, giving an expressing plasmid containing the antisense ACC synthase gene.
出处
《东北农业大学学报》
CAS
CSCD
2006年第1期22-27,共6页
Journal of Northeast Agricultural University
基金
黑龙江省科技攻关项目(GB01B402-01)
关键词
薄皮甜瓜
ACC合成酶基因
反义基因
反义表达载体
Cucumis melo cv. Qitian I
ACC synthase gene, antisense gene, antisense expression vector
