铜绿假单胞菌脂肪酶LipA基因在枯草芽孢杆菌pab02中的整合表达研究
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- 英文论文题名：Cloning and Integrative Expression of Lipase Genes of Pseudomonas aeruginosa in Bacillus subtilis
- 论文作者：张飞燕 ; 何敏 ; 王振华 ; 潘康成 ; 冯杰 ; 唐慧琴
- 英文论文作者：Feiyan Zhang ; Min He ; Zhenhua Wang ; Kangcheng Pan ; Jie Feng ; Huiqin Tang ; Animal Microecology Institute ; College of Veterinary Medicine ; Sichuan Agricultural University ; Deyang Animal Husbandry and Food Bureau ; Chengdu agricultural science and technology Career Academy
- 年：2016
- 作者机构：四川农业大学动物医学院动物微生态研究中心;德阳市畜牧食品局;成都农业科技职业学院;
- 论文关键词：铜绿假单胞菌 ; 脂肪酶 ; 同源重组 ; 整合表达 ; 枯草芽孢杆菌
- 英文论文关键词：Pseudomonas aeruginosa ; lipase ; homologous recombination ; integrative expression ; Bacillus subtilis
- 会议召开时间：2016-11-04
- 会议录名称：中国畜牧兽医学会动物微生态学分会第五届第十二次全国学术研讨会论文集
- 语种：中文
- 分类号：Q936
- 学会代码：ZGXJ
- 会议名称：中国畜牧兽医学会动物微生态学分会第五届第十二次全国学术研讨会
- 会议地点：中国山东青岛
- 主办单位：中国畜牧兽医学会动物微生态学分会
- 学会名称：中国畜牧兽医学会
- 页数：1
- 文件大小：83k
- 原文格式：O
- 会议级别：全国
摘要
旨在对铜绿假单胞菌脂肪酶LipA基因进行克隆,构建整合表达质粒并在枯草芽孢杆菌pab02中整合表达。根据GenBank中报道的铜绿假单胞菌脂肪酶A基因序列设计引物,扩增酶基因并测序分析,构建整合表达质粒pHSG398-16S-1ipA,转化枯草芽孢杆菌pab02并诱导表达。结果,成功从铜绿假单胞菌A05菌株中扩增得到长1092bp脂肪酶A基因,与铜绿假单胞菌脂肪酶A基因(登录号AB290342)同源性高达98%以上;该基因共编码311个氨基酸,包含由26个氨基酸残基组成的信号肽和285个氨基酸残基组成的成熟肽;成功构建了整合表达质粒pHSG398-16S-1ipA并整合到枯草芽孢杆菌pab02染色体上,获得了重组菌pab02-1ipA,SDS-PAGE分析显示脂肪酶蛋白得到了有效表达,分子量约37kDa。结果表明,成功构建了基于枯草芽孢杆菌16S rRNA为同源序列的整合表达质粒pHSG398-16S-1ipA,获得了整合表达重组枯草芽孢杆菌pab02-1ipA。
This experiment was conducted to construct an integrative expression vector and expressions of Pseudomonas aeruginosa lipase A gene in Bacillus subtilis pab02.A pair of primers was designed by Pseudomonas aeruginosa lipase A gene sequence from Genbank().The lipase A gene of P.aeruginosa A05 was cloned and sequenced.The integrated expression plasmid pHSG398-16S-lipA was constructed and transformed into Bacillus subtilis pab02.The recombinant B.subtilis pab02-lipA was induced to expresse lipase.The results showed that the length of lipase gene was 1092 bp ecoding311 amino acids,containing a signal peptide of 26 amino acids and a mature peptide of 285 amino acids.It shared 98%homology with the lipase A gene of P.aeruginosa(Accession No.AB290342).The integrated expression plasmid pHSG398-16S-lipA was constructed successfully and transformed into B.subtilis pab02.SDS-PAGE Showed that the lipase A was successfully expressed and the molecule weight was about 37 KDa.The results indicated that the integration of homologous sequence expression plasmid pHSG398-16s-lipA was constructed successfully and obtained the integrated expression of the recombinant B.subtilis pab02-lipA.
This experiment was conducted to construct an integrative expression vector and expressions of Pseudomonas aeruginosa lipase A gene in Bacillus subtilis pab02.A pair of primers was designed by Pseudomonas aeruginosa lipase A gene sequence from Genbank().The lipase A gene of P.aeruginosa A05 was cloned and sequenced.The integrated expression plasmid pHSG398-16S-lipA was constructed and transformed into Bacillus subtilis pab02.The recombinant B.subtilis pab02-lipA was induced to expresse lipase.The results showed that the length of lipase gene was 1092 bp ecoding311 amino acids,containing a signal peptide of 26 amino acids and a mature peptide of 285 amino acids.It shared 98%homology with the lipase A gene of P.aeruginosa(Accession No.AB290342).The integrated expression plasmid pHSG398-16S-lipA was constructed successfully and transformed into B.subtilis pab02.SDS-PAGE Showed that the lipase A was successfully expressed and the molecule weight was about 37 KDa.The results indicated that the integration of homologous sequence expression plasmid pHSG398-16s-lipA was constructed successfully and obtained the integrated expression of the recombinant B.subtilis pab02-lipA.
引文
[1]张搏,杨江科,闫云君.假单胞菌脂肪酶的研究进展[J].生物技术通报,2007,2:52-57.
[2]PanizzaP,SyfantouN,Pastor F I,etal.Acidic lipase Li PI.3 from a Pseudomonas fluorescens-like strain displays unusual properties and shows activity on secondary alcohols[J].J Appl Microbiol,2013,114(3):722-732.
[2]PanizzaP,SyfantouN,Pastor F I,etal.Acidic lipase Li PI.3 from a Pseudomonas fluorescens-like strain displays unusual properties and shows activity on secondary alcohols[J].J Appl Microbiol,2013,114(3):722-732.