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采用RT-PCR技术,对从美国和埃及引进的两个尼罗罗非鱼Oreochromis niloticus品系(以下分别简称为美国尼罗和埃及尼罗)热休克蛋白70(Heat shock proteins 70,Hsp70)基因进行扩增并克隆测序。获得两个品系罗非鱼Hsp70基因完整编码区(code sequences,CDS)cDNA序列,全长为1 923 bp,编码为640个氨基酸,含有Hsp70家族3个签名序列DLGTTYS、IFDLGGGTFD和VVLVGGSTRIPKIQK,以及C末端保守序列EEVD,核定位信号标签KRKHKKDISQNKRALRR;Dank特征基序DLGTT-S-V;胞质Hsp70特征基序EEVD;靠近C端的GGMP4肽序列;2个糖基化位点NKSI和NVSA。结合BLAST分析结果,可以确认,获得的cDNA序列为美国尼罗和埃及尼罗两个品系罗非鱼的完整编码序列。序列同源性分析表明,美国尼罗和埃及尼罗有3个氨基酸位点和12个核苷酸位点不同,美国尼罗Hsp70基因氨基酸序列与莫桑比克罗非鱼同源性最高为99.7%,而与家鼠同源性最低为83.6%。
The heat shock proteins 70 (Hsp70) genes of two strains of Oreochromis niloticus (hereinafter referred to as Nile and Nile, respectively) introduced from the United States and Egypt were analyzed by RT-PCR Amplify and clone the sequencing. The cDNA sequence of the coding sequence of Hsp70 gene from two strains of tilapia was obtained. The full-length cDNA sequence of the coding sequence of Hsp70 gene was 1 923 bp in length and encoded 640 amino acids. It contained three signature sequences DLGTTYS, IFDLGGGTFD and VVLVGGSTRIPKIQK of Hsp70 family, Conserved sequence EEVD, nuclear localization signal tag KRKHKKDISQNKRALRR; Dank characteristic motif DLGTT-SV; cytoplasmic Hsp70 signature motif EEVD; GGMP4 peptide sequence near C-terminal; two glycosylation sites NKSI and NVSA. Combined with the results of BLAST analysis, it was confirmed that the cDNA sequence obtained was the complete coding sequence of two strains of tilapia, Nile, Egypt and Nile, Egypt. Sequence homology analysis showed that there are three amino acid residues and 12 nucleotide residues in the Nile of the United States and Nile of Egypt. The amino acid sequence of the Nile Hsp70 gene in the United States has the highest homology of 99.7% with the Mozambique tilapia, While the homology with domestic mice is the lowest is 83.6%.