棉蚜catb5基因rnai载体的构建及其对烟草的转化-生物技术通报.pdfVIP

生物技术通报 BIO TE CHN OLOG Y B ULLETIN 20 10 8 Catb5 RNAi 李晓明 高朝宝 彭明 崔百明 ( , 832003) : 依据细胞色素氧化酶亚基 序列的多态性设计特异性引 , 通过 PCR 技术扩增出的特异性条带鉴定为棉蚜 提取棉蚜总 RNA, 利用 RTPCR技术从棉蚜中扩增出大小为 515 bp的C atb5 目的基因, 进一步将两段 Catb5基因分别正向和 反向连接到植 表达载体 pB i35SG 12上, 构建了 pB i35SC5 RNA i表达载体, 并通过农杆菌介导法导入烟草中, PCR检测表明已 得到转基因烟草再生苗 : 棉蚜 C atb5基因 RNA i 表达载体 烟草 Construction of RNAiExpression Vector forAp h is gossipy Catb5 Gene and Transformation of Tobacco LiX iaom ing Gao Chaobao P ngM ing Cu iBami ing (K ey Laboratory of Agriculture B iotechnology of Shihez i University, College of Lfi e Science, Shihezi University, Shihez i 832003) Abstrac:t Aphis gosspi y w as id ntifi d by PCR w ith prmi rs accord ing to th s qu ncing l ngth polymorph ism m thod bas d on th cytochrom e ox i ase subun it I g n . Th total RNA w as xtract d from A. gosspi y and th 515 bp l ngth product of C atb5 g n w as clon d by RTPCR. RNA i v ctor pB i35SC5 w as construct d by ins rting th tw o C atb5 g n s into plant xpr ssion v ctor pB i35SG12 forw ard and r v rs ly. Th v ctor is transform d into tobacco m d iat d byAgrobacterium tum ef aciens GV3101. Transg n ic plants w r scr n d by kanamy cin r sistant and v rifi d by PCR analy sis, w hich provid s a foundation for furth r xp rmi nt of r s isting A. gos spi y. K ey w ords: Aphis gosspi y Catb5 g n RNA i pB i35SC5 T obacco RNA ( RNA int rf r nc , RNA i) , , , mRNA , , RNA D ic r 22 bp , , , RNA ( sm all int rf ring RNAs, siRNA s)siRNAs 1981 Schn pfh [ 1] RNA ( RNAin Bt duc d sil ncing comp l x, RISC), , , siRNAsmRNA,