中国组织工程研究2012，Vol.16Issue(2)：311-314,4.DOI:10.3969/j.issn.1673-8225.2012.02.028
人端粒酶催化亚基反义RNA重组腺病毒载体的构建与鉴定
Construction and identification of recombinant adenovirus vector of human telomerase reverse transcriptase
摘要
Abstract
BACKGROUND: Human telomerase reverse transcriptase (hTERT) closely correlates with c-myc in laryngeal squamous cell carcinoma. OBJECTIVE: To construct recombinant adenovirus vector of hTERT containing c-myc promoter and to observe the effects of expressed hTERT on cell growth. METHODS: Using the method of reverse transcription in artificial synthesis to design the gene synthesis primer and target fragment ashTERT (322 bp) was obtained using overlapping PCR. The target fragment ashTERT was cloned into the vector pUC57(2.7 kb) and then heat-shock transformed into the competent cells E.coli. With Bacteria inspection, PCR, identification of positive clones, plasmid pUC57-ashTERT was extracted and sequenced. The target gene fragments were subcloned into the shuttle vector pshuttle-CMVneo and then the recombinant adenovirus plasmid pAd-ashTERT was constructed. Subsequently, the recombinant adenovirus plasmid pAd-ashTERT was transformed into the 293A cells for package, identification and titer determination of adenovirus. RESULTS AND CONCLUSION: pUC57-ashTERT plasmid was extracted and its sequence corresponded to sequencing requirement. Recombinant adenovirus vector of hTERT with relatively strong ability of infection was successfully constructed after pUC57-ashTERT plasmid was transformed into 293 cells.关键词
人端粒酶催化亚基/瘢痕疙瘩/成纤维细胞/腺病毒/基因治疗分类
医药卫生引用本文复制引用
彭智,贾振华,唐红伟,黄海华,郭晓瑞,韦有万,王香梅,张培华..人端粒酶催化亚基反义RNA重组腺病毒载体的构建与鉴定[J].中国组织工程研究,2012,16(2):311-314,4.基金项目
广东省社会发展项目 (2007B031510001),hTERT调控区反义RNA腺病毒载体的构建及其对瘢痕疙瘩成纤维细胞作用. （2007B031510001）
