农业生物技术学报2013，Vol.21Issue(7)：847-854,8.DOI:10.3969/j.issn.1674-7968.2013.07.011
盐角草Cu/Zn-SOD基因的克隆及耐盐性分析
Cloning and Salt-tolerance Analysis of the Salicornia europaea Gene Cu/Zn-SOD
摘要
Abstract
In order to study the salt-tolerant mechanism of the Salicornia europaea gene Cu/Zn-SOD,conserved sequences of Cu/Zn-SOD from the known plants were designed for degenerate primers in this research.The coding region of Cu/Zn-SOD was amplified by RACE technology.Amino acid sequences and multiple sequences alignment were analyzed by biological software.The gene carried by prokaryotic expression vector pet-duetl was transferred to Escherichia coli BL21 and expressed after IPTG induction.The status of recombinants cultivated in LB medium with different NaCl concentrations were observed.The salttolerant function of Cu/Zn-SOD was assessed by OD600 value.As a result,the Salicornia europaea gene Cu/Zn-SOD was amplified by degenerate primers and RACE technology.The full-length of Cu/Zn-SOD cDNA (GenBank number:JQ074238.2) was 660 bp,containing a 459 bp open reading frame (ORF).The ORF encoded a 152-amino acid polypeptide with a predicted molecular mass of 15.1 kD.The predicted amino acid sequences showed 96％ similarity with the corresponding Suaeda salsa gene sequences and 88％ similarity with the Capsicum chinense gene sequences.Biological software analysis implied that the Cu/Zn-SOD protein existed in cytoplasm.We constructed a prokaryotic expression vector pETCu/Zn-SOD,contrasted by control pETDuet-1,and transformed them into Escherichia coli BL21.Protein was expressed by IPTG induction.The target protein was successfully expressed,as the molecular mass of it accorded with theoretical value by SDS-PAGE electrophoresis.Salt-tolerance analysis showed that recombinant bacteria BL21 (pET-Cu/Zn-SOD) grew significantly better in the high salinity mediums than that of the control bacteria BL21 (pETDuet-1).It indicates that the Salicornia europaea gene Cu/Zn-SOD may play an important role in the salt stress.关键词
盐角草/超氧化物歧化酶/基因克隆/耐盐性Key words
Salicornia europaea/ Superoxide dismutase(SOD)/ Gene cloning/ Salt tolerance引用本文复制引用
臧洁,余梅,王先磊,王荣富,李娟,吉虎,徐凯,鲁茂龙..盐角草Cu/Zn-SOD基因的克隆及耐盐性分析[J].农业生物技术学报,2013,21(7):847-854,8.基金项目
国家"十二五"科技支撑计划课题(No.2011BAD01B01)、安徽农业大学人才引进项目和昆山科腾生物科技有限公司基因工程项目 （No.2011BAD01B01）
