人骨形成蛋白2和人骨形成蛋白7成熟肽编码区基因克隆和真核表达载体的构建  

Construction of Human BMP-2 and-7 Mature Peptide cDNA Expression Plasmids

作　　者：吉建新[1] 廖伟娇[2] 刘利东[2] 卢春生[2] 朱伯平[3] 范婷婷[2] 

机构地区：[1]广州医学院第一附属医院口腔科,广州510120 [2]广州医学院第一附属医院检验科,广州510120 [3]广州医学院第一附属医院输血科,广州510120

出　　处：《广东医学》2010年第4期410-413,共4页Guangdong Medical Journal

基　　金：广东省社会发展领域科技计划项目(编号:2007B060401069);广州市科技局攻关项目(编号:2007Z3-E0191);广州市教育局项目(编号:61105)

摘　　要：目的克隆和构建人骨形成蛋白2(BMP-2)和人骨形成蛋白7(BMP-7)成熟肽真核表达载体。方法采用Trizol法从人骨肉瘤细胞中提取制备总RNA,利用RT-PCR、PCR转录、扩增BMP-2和BMP-7获得成熟肽基因,与真核表达载体pcDNA3.1(+)连接,构建真核表达重组子pcDNA3.1(+)/BMP-2MP和pcDNA3.1(+)/BMP-7MP。结果总RNA提取液电泳可见在28S、18S和5S处出现明显的条带,PCR和酶切可在439bp和364bp出现条带,阳性克隆测序结果与Genebank中登录的序列一致。结论成功克隆出人BMP-2MP和BMP-7MP基因,成熟肽基因已与pcDNA3.1(+)连接,成功构建其真核表达重组子pcDNA3.1/BMP-2MP和pcDNA3.1/BMP-7MP,为进一步研究BMP-2、BMP-7的功能及其成熟肽基因在骨髓间充质细胞(hMSCs)中表达以及在骨组织工程中的应用研究奠定了基础。Objective To clone the mature peptide coding gene of human bone morphogenetic protein-2（BMP-2）or human bone morphogenetic protein-7（BMP-7）for construction of the eukaryotic expression vectors pcDNA3.1（＋）/BMP-2MP and pcDNA3.1（＋）/BMP-7MP.Methods Total RNA was isolated from human osteosarcoma tissue according to Trizol method.Subsequently,the mature peptides coding genes of human BMP-2 and BMP-7 were obtained through reverse transcription PCR,PCR transcribed,amplified and recombined with the eukaryotic expression plasmid pcDNA3.1（＋）.Results The recombined vectors were proved by electrophoresis,PCR-RFLP and cDNA sequencing matching with Genebank.Conclusion The eukaryotic expression vectors of pcDNA3.1（＋）/BMP-2MP and pcDNA3.1（＋）/BMP-7MP were successfully constructed.

关 键 词：骨形成蛋白-2 骨形成蛋白-7 基因克隆 真核表达载体 

分 类 号：R329.26[医药卫生—人体解剖和组织胚胎学]