赤拟谷盗HSP70基因克隆和竞争定量PCR检测

为研究赤拟谷盗Tribolium castaneum受到热胁迫后高度保守的热激蛋白70（heat shock protein 70，HSP70）基因的表达变化，本研究扩增了681 bp的赤拟谷盗hsp70片段，编码227个氨基酸残基，GenBank登录号为HM345948。同源性分析表明：赤拟谷盗hsp70核苷酸序列与马铃薯甲虫Leptinotarsa decemlineata的hsp70（GenBank登录号：AF322911.1）同源性最高，为97%；其推测的蛋白序列与马铃薯甲虫、甘蓝夜蛾Mamestra brassicae、黑腹果蝇Drosophila melanogaster和美洲斑潜蝇Liriomyza sativae的HSP70蛋白均有94%以上的同源性。利用RT-PCR技术得到与赤拟谷盗hsp70进行竞争定量的内部竞争物， 以等量的目标cDNA和一系列稀释的竞争模板进行竞争PCR扩增，构建了hsp70的竞争定量PCR检测体系， 该体系标准曲线的线性方程为Y=1.032X-1.618 (r²=0.975)。这些结果为赤拟谷盗的hsp70定量检测提供了方便，并为热控技术防治害虫提供了基础资料。

Cloning and quantitative competitive PCR assay of HSP70 gene in Tribolium castaneum(Coleoptera:Tenebrionidae)

To study the expression changes of heat shock protein 70(a highly conserved protein)in Tribolium castaneum after exposure to heat stress,a fragment of 681 bp encoding for HSP70 was amplified and sequenced from T.castaneum.The fragment encoded 227 amino acid residues with the GenBank accession no.HM345948.The result of homology analysis showed that this fragment shared 97% identity with hsp70 from L.decemlineata(AF322911.1).Comparison of the deduced amino acid sequences of HSP70 in T.castaneum with those in ...