人内皮细胞抑制素基因及其表达产物的生产和应用

摘要

本发明属于基因工程技术领域。本发明以人胶原蛋白XⅧcDNA为模板,经PCR方法合成人内皮细胞抑制素基因,并构建了含有该基因的载体和重组菌株。特别是含有该基因的大肠杆菌表达载体pBVE20及其转化的大肠杆菌DH5α(pBVE20)重组株和利用此菌株生产重组人内皮细胞抑制素的方法。此菌株具有表达量高,表达产物易纯化等优点。本发明将为获得来源稳定价格便宜的重组人内皮细菌抑制素并用其制备新一代抗癌药物提供一条切实可行的途径。

主权项

1．人内皮细胞抑制素基因，其特征是：它是以人胶原蛋白ⅩⅧ cDNA为模板，经PCR方法扩增而得到的来源于人胶原蛋白ⅩⅧ cDNA的3’端所对应的基因片段，它是人胶原蛋白ⅩⅧ cDNA上从终止密码算起至上游共184个密码子的DNA序列，包括起始密码ATG共555bp的片段，其DNA的核苷酸序列及其所编码的氨基酸序列如下：ATG CAC AGC CAC CGC GAC TTC CAG CCG GTG CTC CAC CTG GTT GCG CTC 48Met His Ser His Arg Asp Phe Gln Pro Val Leu His Leu Val Ala LeuAAC AGC CCC CTG TCA GGC GGC ATG CGG GGC ATC CGC GGG GCC GAC TTC 96Asn Ser Pro Leu Ser Gly Gly Met Arg Gly Ile Arg Gly Ala Asp PheCAG TGC TTC CAG CAG GCG CGG GCC GTG GGG CTG GCG GGC ACC TTC CGC 144Gln Cys Phe Gln Gln Ala Arg Ala Val Gly Leu Ala Gly Thr Phe ArgGCC TTC CTG TCC TCG CGC CTG CAG GAC CTG TAC AGC ATC GTG CGC CGT 192Ala Phe Leu Ser Ser Arg Leu Gln Asp Leu Tyr Ser Ile Val Arg ArgGCC GAC CGC GCA GCC GTG CCC ATC GTC AAC CTC AAG GAC GAG CTG CTG 240Ala Asp Arg Ala Ala Val Pro Ile Val Asn Leu Lys Asp Glu Leu LeuTTT CCC AGC TGG GAG GCT CTG TTC TCA GGC TCT GAG GGT CCG CTG AAG 288Phe Pro Ser Trp Glu Ala Leu Phe Ser Gly Ser Glu Gly Pro Leu LysCCC GGG GCA CGC ATC TTC TCC TTT GAC GGC AAG GAC GTC CTG AGG CAC 336Pro Gly Ala Arg Ile Phe Ser Phe Asp Gly Lys Asp Val Leu Arg HisCCC ACC TGG CCC CAG AAG AGC GTG TGG CAT GGC TCG GAC CCC AAC GGG 384Pro Thr Trp Pro Gln Lys Ser Val Trp His Gly Ser Asp Pro Ash GlyCGC AGG CTG ACC GAG AGC TAC TGT GAG ACG TGG CGG ACG GAG GCT CCC 432Arg Arg Leu Thr Glu Ser Tyr Cys Glu Thr Trp Arg Thr Glu Ala ProTCG GCC ACG GGC CAG GCC TCC TCG CTG CTG GGG GGC AGG CTC CTG GGG 480Ser Ala Thr Gly Gln Ala Ser Ser Leu Leu Gly Gly Arg Leu Leu GlyCAG AGT GCC GCG AGC TGC CAT CAC GCC TAC ATC GTG CTC TGC ATT GAG 528Gln Ser Ala Ala Ser Cys His His Ala Tyr Ile Val Leu Cys Ile GluAAC AGC TTC ATG ACT GCC TCC AAG TAG 555Asn Ser Phe Met Thr Ala Ser Lys stop