以温和噬菌体ρ11为载体克隆α-淀粉酶基因

CLONING OF α-AMYLASE GENE WITH THE TEMPERATE PHAGE ρ11

温和噬菌体 p11 DNA 和枯草芽孢杆菌(Bacillus subtilis)168染色体 DNA 用 BamH1酶消化和 T4 DNA 连接酶连接,转化到被噬菌体ρ11溶源化了的501菌株中,经过体内交换整入到染色体上,培养48小时后,从1020个菌落中挑出含 Arol^+-Amy^+基因的3个转化子,分别命名为ρ11Amy1、ρ11Amy2和ρ11Amy3。取ρ11Amy1 进行繁殖和丝裂霉素 C 诱导,诱导后的重组噬菌体用于测定噬斑形成能力和转导活性,提取其 DNA 再用于转化,均证明构建成了含淀粉酶基因的重组特异性转导噬菌体。

DNA from a temperate phage ρll andchromosomal DNA of Bacillus subtilis 168were digested with endonuclease BamHI andthen ligated with T4 ligase.The ligated DNAfragments were used to transform a lysogenicstrain B.subtilis 501.After incubation for48h,3 Arol^+-Amy^+ transformants were se-lected from 1020 colonies.They were tenta-tively named ρllAmyl,ρllAmy2 and ρll-Amy3.ρll Amyl was grown and treated withmitomycin C.The induced phages weretested for abilities to form plaques and todo transducing activity.Extracted DNAsfrom ρllAmyl were used to trasform theamy- recipient cells.The results shown thatspecial phage was constructed.