马铃薯甲虫热激蛋白基因hsp60、hsp70和hsp90的克隆及温度胁迫下的表达-cloning and expression of hsp 60, hsp 70 and hsp 90 genes of potato beetle under temperature stress.docxVIP

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马铃薯甲虫热激蛋白基因hsp60、hsp70和hsp90的克隆及温度胁迫下的表达-cloning and expression of hsp 60, hsp 70 and hsp 90 genes of potato beetle under temperature stress.docx

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马铃薯甲虫热激蛋白基因hsp60、hsp70和hsp90的克隆及温度胁迫下的表达-cloning and expression of hsp 60, hsp 70 and hsp 90 genes of potato beetle under temperature stress

青海大学青海大学硕士学位论文 Abstract II II Abstract As one of international major quarantine pest, Colorado potato beetle, Leptinotarsa decemlineata, has strong ability of stress resistance, especially for the extreme temperature stress resistance. The overall goal of this study is to better understand the molecular mechanism of temperature stress resistance in L. decemlineata. The full-length cDNA encoded the HSP60, HSP70 and HSP90 from L. decemlineata were cloned analysed by Bioinformations methods, RT-PCR and RACE approaches. The relative expression levels of Ld-hsp90 and Ld-hsp60 in adults L. decemlineata under a series of temperatures were detected by Real -Time PCR. The main results are as follows: This was the first time that full-length cDNA sequences encoded HSP90 and HSP60 from L. decemlineata were cloned, which were named to Ld-hsp90 and Ld-hsp60 respectively. The complete length cDNA were 2489bp and 2234bp respectively, the length of 5 untranslated region (UTR) were 113bp and 101bp respectively, and the length of 5 UTR were 216bp and 402bp respectively. The complete ORF (2160bp, 1731bp) encodes a 719 and a 576 amino acid protein respectively. The amino acid sequences contain classi cal and highly conserved signature sequences of HSP90 and HSP60 family respectively. Meanwhile, 3 full-length cDNA sequences of encoded HSP70 were cloned in L. decemlineata, which were named to Ld-hsp70a (2199bp), Ld-hsp70b (2397) and Ld-hsp70c (2482bp) respectively. The length of 5 UTR were 92bp, 132bp and 289bp respectively, and 3 UTR were 160bp, 291bp and 203bp. The complete ORF (1947bp, 1974bp and 1890bp) encodes a 648, a 657 and a 629 amino acid protein respectively. The 3 amino acid sequences of HSP70 share a 71.50% identity, and they all contain the classical and highly conserved signature sequences of HSP70 family. III III The Real-Time PCR results indicated that the relative expression levels of Ld-hsp90 was significantly increased after 1 hour exposed t o 38℃ and 45℃ when comp