拟穴青蟹PDI保守区域的克隆与序列分析

蛋白二硫键异构酶(PDI)是真核生物重要的多功能蛋白,其基因结构在虾蟹类尚未报道.从拟穴青蟹Scylla pa-ramamosain(Estampador,1949)脑组织中分离纯化总RNA,经逆转录得到cDNA第一条链,并以之为模板利用RACE(rapid-amplification of cDNA ends)技术,扩增出一条950 bp的cDNA.将所得cDNA克隆到质粒载体pTZ57R/T,转化大肠杆菌DH5α细胞并对筛选的阳性克隆测序.通过与目前数据库中序列的比较,在此cDNA编码蛋白中发现其中101个氨基酸与其他物种已知蛋白二硫键异构酶(PDI)中的PDI_a_PDI_a'_C保守区域相似率在64%～79%之间.确定此cDNA编码拟穴青蟹PDI_a_PDI_a'_C保守区域.

Cloning and Sequence Analysis of Scylla paramamosain Protein-disulfide Isomerase Conserved Domain

The protein disulfide isomerase severs to be a muhifunctional protein of great importance to the eukaryotes.The total RNAs isolated from the brain of Scylla paramamosainn(Estampador,1949)were reverse transcribed to the first strands of cDNA.A cDNA fragment of 950 bp was amplified from the first strands of cDNA by RACE(rapid-amplification of cDNA ends)method.This fragment was inserted into plasmid pTZ57R/T for screening and DNA sequencing.Compared with current sequences in the data-base,a 101-aa domain,which was found in the putative protein,had 64%～79%homology with the PDI_a_PDI_a'_C conserved do-mains of protein-disulfide isomerases(PDIs)in other species.This cDNA was supposed to encode Scylla paramamosain PDI_a_PDI_a'_C conserved domain.