CAJ | 학술논문

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28 S、18 S和5 S处出现明显的条带,PCR和酶切可在439 bp和364 bp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.
목적 극륭화구건인골형성단백2(BMP-2)화인골형성단백7(BMP-7)성숙태진핵표체재체.방법 채용Trizol법종인골육류세포중제취제비총RNA,이용RT-PCR、PCR전록、확증BMP-2화BMP-7획득성숙태기인,여진핵표체재체pcDNA3.1(+)련접,구건진핵표체중조자pcDNA3.1(+)/BMP-2MP화pcDNA3.1(+)/BMP-7MP.결과 총RNA제취액전영가견재28 S、18 S화5 S처출현명현적조대,PCR화매절가재439 bp화364 bp출현조대,양성극륭측서결과여Genebank중등록적서렬일치.결론 성공극륭출인BMP-2MP화BMP-7MP기인,성숙태기인이여pcDNA3.1(+)련접,성공구건기진핵표체중조자pcDNA3.1/BMP-2MP화pcDNA3.1/BMP-7MP,위진일보연구BMP-2、BMP-7적공능급기성숙태기인재골수간충질세포(hMSCs)중표체이급재골조직공정중적응용연구전정료기출.
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.