CAJ | 학술논문

背景:转化生长因子β1能促进多种细胞的生长增殖,是调控骨髓间充质干细胞向成骨方向定向分化的主要生长因子.目的:利用AdMax系统构建人转化生长因子β1(hTGF-β1)基因的重组腺病毒表达载体.方法:采用PCR方法克隆人转化生长因子β1基因cDNA后,插入线性化表达载体pAV-MCMV-EGFP-3FLAG中,转化E.coli DH5α感受态细胞,构建pAV-MCMV-hTGF-β1重组质粒.结果与结论:含目的基因的重组腺病毒质粒共转染293细胞进行病毒包装、扩增后,经PCR、Western Blot检测得到转化生长因子β1重组腺病毒,其滴度约为1.25×1010 pfu/mL.表明利用AdMax系统成功可构建人转化生长因子β1腺病毒载体,并可满足进一步的转化生长因子β1成骨作用的研究.
배경:전화생장인자β1능촉진다충세포적생장증식,시조공골수간충질간세포향성골방향정향분화적주요생장인자.목적:이용AdMax계통구건인전화생장인자β1(hTGF-β1)기인적중조선병독표체재체.방법:채용PCR방법극륭인전화생장인자β1기인cDNA후,삽입선성화표체재체pAV-MCMV-EGFP-3FLAG중,전화E.coli DH5α감수태세포,구건pAV-MCMV-hTGF-β1중조질립.결과여결론:함목적기인적중조선병독질립공전염293세포진행병독포장、확증후,경PCR、Western Blot검측득도전화생장인자β1중조선병독,기적도약위1.25×1010 pfu/mL.표명이용AdMax계통성공가구건인전화생장인자β1선병독재체,병가만족진일보적전화생장인자β1성골작용적연구.
BACKGROUND:Transforming growth factor-beta1 (TGF-β1) can promote the growth and proliferation of many cel s and is one of main growth factors that regulate osteoblast differentiation of bone marrow mesenchymal stem cel s. OBJECTIVE:To construct a recombinant adenoviral vector carrying the human TGF-β1 gene using the AdMax system which may be applicable in gene therapy of bone defect. METHODS:The hTGF-β1 gene sequence was amplified by PCR from cDNA template and then was inserted into shuttle plasmid pAV-MCMV-EGFP-3FLAG. The recombinant shuttle plasmid was constructed and transformed into E.coli DH5α.The recombinant pAV-MCMV-hTGF-β1 was obtained. RESULTS AND CONCLUSION:The recombinant pAV-MCMV-hTGF-β1 was packaged and amplified in 293 cel s. Then the recombinant TGF-β1 adenovirus was constructed successful y. After measuring the titre of virus (1.25×1010 pfu/mL), the target gene was evaluated by PCR and Western Blot. These findings suggest that construction of hTGF-β1 adenoviral vector was successful y achieved using the AdMax system and may be available in the future study of TGF-β1 gene therapy.