中文摘要：

本研究基于一株对小菜蛾（Plutella xylostella）幼虫具有高毒力的苏云金芽胞杆菌（Bacillus thuringiensis）菌株FH21,利用改进的限制性片段长度多态性聚合酶链反应（polymerase chain reactionrestriction fragment length polymorphism,PCR-RFLP）,从FH21中成功克隆出杀虫晶体蛋白（insecticidal crystal proteins）基因cry1Bb3、cry1Da4、cry1Ja3和cry1Id3,Gen Bank登录号分别为KJ619659、KJ619660、JQ228425和KJ619661。将上述4个基因插入表达载体p EB,转化大肠杆菌（Escherichia coli）菌株Rosetta（DE3）,在低温条件下经异丙基-β-D-硫代半乳糖苷（isopropylβ-D-1-thiogalactopyranoside,IPTG）诱导表达。FH21总蛋白与4种蛋白的表达产物采用饲喂法分别对小菜蛾、棉铃虫（Helicoverpa armigera）和亚洲玉米螟（Ostrinia furnacalis）进行杀虫活性测定。结果表明,FH21总蛋白对3种昆虫均有较好的杀虫活性,致死中浓度（lethal concentration 50,LC50）分别为0.801、0.502和2.858 mg/L。Cry1Bb3、Cry1Ja3和Cry1Id3蛋白对小菜蛾和亚洲玉米螟有较好的杀虫活性,LC50分别为1.860、1.697、1.538、1.800、2.270和0.840 mg/L。本研究表明,Bt FH21菌株对多种农业害虫具有很高的杀虫活性,同时对其进行杀虫基因测定并对杀虫基因进行了杀虫活性测定,几种cry1类基因的发现为重要农业害虫的防治提供了更多的选择。

英文摘要：

This study was based on Bacillus thuringiensis（Bt） strain FH21 with high toxic of Plutella xylostella larvae. Using an improved polymerase chain reaction- restriction fragment length polymorphism（PCR- RFLP） method, cry1Bb3, cry1Da4, cry1Ja3 and cry1Id3 genes were cloned from strain FH21 with the Gen Bank No. KJ619659, KJ619660, JQ228425 and KJ619661, respectively. The 4 genes were inserted into expression vector p EB, and transformed into Escherichia coli strain Rosetta（DE3）, which were induced by isopropyl β- D- 1- thiogalactopyranoside（IPTG） at low temperature. The total protein of FH21 and the 4proteins were used for bioassay on P. xylostella, Helicoverpa armigera and Ostrinia furnacalis. The total protein of FH21 had high insecticidal activity against 3 insects with lethal concentration 50（LC50） of 0.801, 0.502 and2.858 mg/L, respectively. Cry1Bb3, Cry1Ja3 and Cry1Id3 had high insecticidal activity against P. xylostella and O. furnacalis with LC50 of 1.860, 1.697, 1.538, 1.800, 2.270 and 0.840 mg/L, respectively. This study showed that Bt FH21 strain against agricultural pests with a high insecticidal activity, insecticidal genes were assayed and insecticidal activity were determined simultaneously. The cry1 genes provide more choices for important agricultural pest.