小麦条锈菌hsp70基因的克隆及热胁迫下的表达特征分析  

Cloning of a heat shock protein gene hsp70 of Puccinia striiformis f.sp.tritici and its expression in response to high-temperature stress

作　　者：曹华宁 刘博[1] 刘太国[1] 高利[1] 陈万权[1] 

机构地区：[1]中国农业科学院植物保护研究所,植物病虫害生物学国家重点实验室,北京100193

出　　处：《植物保护》2015年第3期19-24,共6页Plant Protection

基　　金：国家重点基础研究发展计划(“973”计划)(2013CB127704);国家自然科学基金项目(31100110);国家科技支撑计划(2012BAS19B04);国家小麦产业技术体系岗位专家专项经费(CARS-03-04B)

摘　　要：为明确热激蛋白70在小麦条锈菌对高温适应性中的作用,采用RACE和PCR技术扩增得到小麦条锈菌hsp70的基因全长,并利用实时荧光定量PCR技术对热胁迫下不同温度敏感类型小麦条锈菌中hsp70的表达特征进行分析。克隆得到的小麦条锈菌hsp70基因组序列全长为2 817bp,包含7个内含子,cDNA序列全长为2 267bp,其中开放阅读框为1 917bp,编码639个氨基酸,分子量为66.7ku,等电点为5.15。编码蛋白含3个保守的HSP70氨基酸序列。qRT-PCR试验结果表明,28℃热激后,hsp70的转录水平随热激时间的延长而略有增加,2h达到最高值。不同温度敏感类型菌株在热激2h后hsp70的相对表达量具有显著差异。低敏感类型菌株hsp70平均表达量为未经处理对照的7.12倍,而高敏感类型菌株hsp70平均表达量仅为对照的1.63倍。To determine the response of Puccinia striiformis f. sp. tritici heat shock protein 70 to high-temperature stress, the Pst hspTO gene was cloned via RACE and PCR, and mRNA expression of hsp70 was determined in represent- ative isolates with low-and high-temperature sensitivity during the heat shock process. The whole genomic DNA se- quence of hspT0 was 2 817 bp, including 7 introns, and the cDNA sequence was 2 267 bp, including an opening reading frame of 1 917 nucleotides. The ORF encodes a HSP70 protein of 639 amino acids, which had a theoretical pI/MW of 5.15/66.7 ku and contained three HSP70 family signature domains. Following heat shock at 28 ℃, expression of hsp70 transcripts was up-regulated in the isolates with high-and low-temperature sensitivity, and reached the maximum level 2 hours after treatment. However, the average expression level was increased by 7.12-fold upon heat shock for 2 h in the isolates with low-temperature sensitivity, but only 1.63-fold in the isolates with high-temperature sensitivity.

关 键 词：小麦条锈菌 温度敏感类型 热激蛋白70 克隆 荧光定量PCR 

分 类 号：S435.121.42[农业科学—农业昆虫与害虫防治]