人EGF受体胞外域cDNA克隆及其序列分析  

Molecular cloning and sequence analysis of cDNA encoding human EGF receptor extracellular domain

作　　者：徐丽慧[1,2] 洪岸[1,2] 何贤辉[3] 

机构地区：[1]暨南大学生物工程研究所 [2].教育部基因组药物工程研究中心广东广州510632 [3]暨南大学组织移植与免疫教育部重点实验室

出　　处：《暨南大学学报（自然科学与医学版）》2005年第2期168-173,179,共7页Journal of Jinan University(Natural Science & Medicine Edition)

基　　金：国家"十五"重大专项基金 (2 0 0 2AA2Z3344 );广东省"十五"重大专项基金 (2 0 0 1A10 90 2 0 8)资助项目

摘　　要：目的:克隆编码人表皮生长因子受体(EGFR)胞外域的cDNA ,构建可溶性EGFR(sEGFR)的哺乳动物细胞表达载体。方法:以RT -PCR方法从人胎盘中克隆编码EGFR胞外域的cDNA ,测定其序列,构建sEGFR真核表达载体。结果:从人胎盘绒毛组织中成功克隆编码EGFR胞外域的cNDA ,通过PCR方法在起始位点前加入Kozak序列并添加终止密码,构建了sEGFR的真核表达载体。该基因编码的蛋白质包含信号肽以及L1、S1、L2和S2等4个结构域。序列分析表明该基因有3个碱基与以往报道的基因不同,即15 6 2G→A、16 2 0G→C、1887T→A ,前两个改变导致氨基酸Arg4 97Lys和Lys5 16Asn的改变,后一个为同义突变Thr6 0 5。结论:成功克隆编码EGFR胞外域的cD NA 。Aim: To clone the cDNA of human EGFR extracellular domain and to construct the mammalian cell expression vector for soluble EGFR (sEGFR). Methods: The cDNA encoding EGFR extracellular domain was cloned by RT-PCR from human placenta and its sequence was determined. The DNA fragment was inserted into pEGFP-N1 to construct an expression vector for sEGFR. Results: The cDNA encoding the extracellular domain of EGFR was cloned from human placenta villi and the expression vector for sEGFR was constructed by PCR using a primer pair to introduce a Kozak sequence before ATG and to insert a stop codon. The protein encoded by this gene contained domains L1, S1, L2, S2 as well as signal peptide. Sequence analysis revealed that there were 3 nucleotides different from the previously published gene, with G1562→A, G1620→C and T1887→A, respectively. The former two nucleotide substitutions led to amino acid changes at position 497 and 516 from Arg and Lys to Lys and Asn respectively, whereas the latter one was a synonymous mutation (Thr605). Conclusion: The cDNA encoding the extracellular domain of EGFR was successfully cloned and the expression vector for sEGFR was constructed.

关 键 词：表皮生长因子受体 可溶性EGFR 单核苷酸多态性 

分 类 号：Q78[生物学—分子生物学]