叶用莴苣热激蛋白LsHsp70-3701基因的克隆及高温胁迫下的表达分析  

Cloning and Expression Analysis of Heat-Shock-Protein Ls Hsp70- 3701 Gene From Leaf Lettuce Under High Temperature Stress

作　　者：赵真真[1] 韩莹琰[1] 范双喜[1] 刘超杰[1] 郝敬虹[1] 李婷[1] 李雅博[1] 

机构地区：[1]北京农学院植物科学技术学院,北京102206

出　　处：《核农学报》2016年第6期1083-1090,共8页Journal of Nuclear Agricultural Sciences

基　　金：国家自然科学基金(31372057;31401883);北京市叶类蔬菜创新团队建设专项(blvt-02);北京市农业科技项目-生菜耐热新品种繁育及安全生产技术试验示范项目(20140134)

摘　　要：为探讨热激蛋白Hsp70基因在高温胁迫下的响应机制及其分子机理,分析Hsp70基因与叶用莴苣耐热性的关系,本研究通过同源克隆及RACE技术,克隆了叶用莴苣热激蛋白Ls Hsp70-3701基因的c DNA全长序列,利用实时荧光定量PCR(qRT-PCR)分析该基因在叶用莴苣热敏品种P-S11和耐热品种G-S59中高温胁迫下的表达差异。该基因c DNA全长为2 191 bp,开放阅读框为1 950 bp,编码649个氨基酸,具有Hsp70家族特有的标签序列,与拟南芥、番茄、小麦等物种的Hsp70同源性达到90%以上。qRT-PCR分析显示,高温胁迫下该基因在2个品种中的表达均上调,在耐热品种中总体表达水平显著高于热敏品种,且在42℃高温下热敏品种P-S11中基因表达随着胁迫时间的增加受到抑制,而耐热品种G-S59中则能长时间保持较高的表达水平。结果表明,Ls Hsp70-3701基因可能与叶用莴苣耐热性相关。本研究为有效解决叶用莴苣高温抽薹问题提供了数据支持。The research aimed at finding out the response mechanisms and molecular mechanisms of heat-shock-protein 70 gene under high temperature stress, then analyzing the relationships between Hsp70 gene and thermo-tolerance of leaf lettuce （Lactuca sativa）. By homologous cloning and RACE technology, the full-length cDNA of LsHsp70-3701 was obtained. Real-time fluorescence quantitative PCR （qRT-PCR） was used to analyze the expression differences between heat-sensitive variety P-S11 and heat-tolerant variety G-S59 under high temperature. LsHsp70-3701 had a cDNA sequence with 2 191 bp in length, which contains a 1 950 bp open reading flame （ORF） and encodes a deduced 649 amino acids. It has the unique tag sequences of Hsp70 family, and it shares 90% homology with other Hsp70 proteins, such as Arabidopsis thaliana （NP_187864.1）, Solanum lycopersicum （NP_001266213.1） and Triticum aestivum （AAB99745.1）. The Real-time PCR results showed that the overall expression level of LsHsp70-3701 in the heat resistant variety was significantly higher than that in the heat sensitive variety although it was upregulated in both varieties. The expression in the heat sensitive variety P-S11 was constrained after a period under 42 ℃, while it can maintain a high level for a long time in the heat resistant variety G-S59. These results indicate that LsHsp70-3701 gene may associat with heat resistance in leaf lettuce. It provided data support for solving the problem of Lactuca sativa prone to bolting under high temperature.

关 键 词：叶用莴苣 HSP70 基因克隆 高温胁迫 表达分析 

分 类 号：S636.2[农业科学—蔬菜学]