苏云金芽胞杆菌新菌株S184crylAb基因的克隆  

Cloning of crylAb Gene from Novel Strain S184 of Bacillus thuringiensis

作　　者：李建华[1] 余健秀[1] 庞义[1] 邓日强[1] 

机构地区：[1]中山大学生物防治国家重点实验室,广州510275

出　　处：《农业生物技术学报》1999年第4期353-357,共5页Journal of Agricultural Biotechnology

基　　金：广东省自然科学基金!963056;980296

摘　　要：通过对GenBank中cryl类基因序列的保守区进行分析，设计一对针对其保守区的引物Y5－1（AGGACCAGGATTTACAGGAGG）和Y3－1（GCTGTGACACGAAGGAIAJAGCCAC），对苏云金芽胞杆菌（Bacillusthuringiensis,Bt）新菌株S184质粒DNA进行扩增，得到一大小为1．5kb的DNA片段，序列分析显示该片段与Cryl类基因高度同源。以此片段为探针，对S184质粒DNA酶切片段进行Southem杂交，在SalⅠ、SacⅠ、SacⅠ＋SalⅠ限制性酶切片段5kb处均只有唯一的阳性信号。将SacⅠ5kb片段克隆至pBluescriptM13-上得到重组质粒pB1Ab，然后进行亚克隆及部分序列测定，结果表明所克隆的片段含有cry1Ab亚类基因。Employing comparer analysis of cryl gene conserved regions, a pair of primers Y5-1 (AGGACCA-GGATTTAGGAGG) and Y3-1 (GCTGTGACACGA A GGATAT AGCCAC) were designed. With the primers,PCR was performed using plasmid DNA from novel strain S184 of Bacillus thuringiensis (Bt) as the template and 1 .5 kb fragment was obtained. The 824-bp 3-end of this PCR product was sequenced, and the homology comparing with crylA gene is up to 99 %. The PCR product was labeled as the probe to hybridize the DNA fragments, which were generated by cleaving the plasmid of strain S184 with several restriction endonuclease. The positive SacI fragment in sise ofabout 5 kb was cloned into pBluescriPt M 13-to create a recombinant plasmid pB 1Ab. Further subcloning and sequencing was cdrried out, revealing that the 5 kb fragment contained a new cry1Ab gene.

关 键 词：苏云金芽孢杆菌 cry1A基因 克隆 菌株 

分 类 号：Q939.124[生物学—微生物学] S482.3[农业科学—农药学]