摘要
【目的】克隆分析菜粉蝶(Pieris rapae)热激蛋白20基因(HSP20),并检测其在温度胁迫下的表达情况,为探析菜粉蝶适应温度胁迫机制提供参考。【方法】以菜粉蝶为试验材料,采用RT-PCR和RACE克隆获得菜粉蝶HSP20基因cDNA全长,并通过生物学软件分析其序列,明确其基因结构特征和亲缘关系;采用实时荧光定量PCR检测菜粉蝶HSP20基因在低温(-20～0℃)和高温(5～45℃)胁迫下的表达响应。【结果】菜粉蝶HSP20基因全长725 bp,5'-端非编码区105 bp,3'-端非编码区109 bp,开放阅读框531 bp,编码176个氨基酸,预测蛋白分子量19.5 kD,理论等电点6.61;其编码氨基酸序列中含有1个典型的HSP20家族结构域(位于第60～142位氨基酸)、1个α-晶体蛋白(位于第60～157位氨基酸)和1个Allato allatostatin(位于第14～24位氨基酸);基序列相似度分析其与二化螟、家蚕和甘蓝夜蛾HSP20蛋白氨基酸相似度较高并聚为一族。该基因随环境温度的升高或降低发生变化,在-5和40℃诱发该基因高效表达。【结论】低温和高温均能诱导菜粉蝶HSP20基因高效表达。
【Objective】Heat shock protein 20 gene(HSP20)from Pieris rapae was cloned and its expression under temperature stress was detected.The result was helpful for further investigation of the adaptation mechanisms of P.rapae to high and low temperature stresses.【Method】Full length cDNA of HSP20 gene of P.rapae was cloned by RT-PCR and RACE with P.rapae as materials.Its sequence was analyzed by biology software,and its gene structural sequence and genetic relationship were also studied.Response of HSP20 gene to low(-20-0℃)and high(5-45℃)temperatures were analyzed by fluorescence quantitative PCR.【Result】The full-length of P.rapae HSP20 was 725 bp,and contained 105 bp 5'-noncoding region,109 bp 3'-noncoding region,a 531 bp open reading frame(ORF),encoded 176 amino acids,with an estimated molecular mass of 19.5 kD and an theoretical isoelectric point(pI)of 6.61.In the encoded amino acid sequences,there was one typical HSP20 family domain(located at 60-142 amino acids),oneα-crystallin(located at 60-157 amino acids)and a Allato allatostatin(located at 14-24 amino acids).The sequence similarities between P.rapae HSP20 and that of Chilo suppressalis,Bombyx mori,Mamestra brassicae were high and they were clustered into one group.The HSP20 expressions changed with temperature,and high expressions of this gene was observed at-5 and 40℃.【Conclusion】Low and high temperatures can lead to the high expression of P.rapae HSP20.
作者
袁远
张连根
高熹
吴国星
张慧
龙华
朱家位
YUAN Yuan;ZHANG Lian-gen;GAO Xi;WU Guo-xing;ZHANG Hui;LONG Hua;ZHU Jia-wei(Yunnan Agricultural University,Kunming 650201,China)
出处
《南方农业学报》
CAS
CSCD
北大核心
2019年第2期286-291,共6页
Journal of Southern Agriculture
基金
云南省重点研发计划项目(2017ZF004)
关键词
菜粉蝶
热激蛋白
HSP20
基因克隆
序列分析
温度胁迫
表达分析
Pieris rapae
heat shock protein
HSP20
gene cloning
sequence analysis
temperature stress
expression analysis
