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热休克蛋白70是已知热休克蛋白家族中最重要的一种,它在细胞内的大量表达可以明显改善细胞的生存能力,提高对环境胁迫的耐受性。为探讨热胁迫对二化螟Chilo suppressa lis幼虫热休克蛋白70表达的影响,采用RT-PCR及RACE技术从二化螟血淋巴细胞中克隆了热休克蛋白70基因全长cDNA序列。该基因全长2102bp,开放阅读框(open reading frame,ORF)为1959bp,编码652个氨基酸;5′非编码区(untranslated region,UTR)为81bp,3′UTR为62bp。从该基因推导的氨基酸序列与其他昆虫的同源序列比较有很高的相似性(73%～97%)。实时定量PCR显示二化螟HSP70基因能被热胁迫诱导表达,幼虫血淋巴细胞的HSP70基因在36℃时表达量最高。流式细胞术研究发现HSP70在蛋白质水平上的表达变化与在mRNA水平上高度一致,说明二化螟HSP70基因在转录及翻译水平上受到热应激的调节。
Heat shock protein 70 is one of the most important heat shock protein family and its large expression in the cell can significantly improve cell viability and increase tolerance to environmental stress. To investigate the effect of heat stress on heat shock protein 70 expression in Chilo suppressa larvae, the full-length cDNA sequence of heat shock protein 70 (HSP70) was cloned from hemolymph of Chilo suppressalis larvae by RT-PCR and RACE techniques. The gene was 2102bp in length and 1959bp in open reading frame (ORF), encoding 652 amino acids. The 5 ’untranslated region (UTR) was 81bp and the 3’UTR was 62bp. The amino acid sequence deduced from this gene has high similarity (73% ~ 97%) with the homologous sequences of other insects. Real-time quantitative PCR showed that HSP70 gene of Chilo suppressalis was induced by heat stress, and HSP70 gene of larval blood lymphocyte was the highest at 36 ℃. Flow cytometry showed that the expression of HSP70 at the protein level was highly consistent with the mRNA level, indicating that HSP70 gene was regulated by heat stress at transcription and translation level.