宫颈癌组织中转录因子skn-基因检测-dna测序.docVIP

宫颈癌组织中转录因子Skn作者：余勐 热西旦 马彩玲， 张富春【关键词】? 宫颈癌 　　摘要: 目的： 从宫颈癌组织中扩增与上皮细胞分化和人乳头瘤病毒(HPV)基因表达相关的转录因子Skn1a cDNA，进而在真核细胞中表达该转录因子以了解其功能。方法：从宫颈癌组织中提取总RNA，以此为模板，RTPCR扩增Skn1a cDNA片段，将该cDNA 片段克隆至pMD18T载体并进行序列分析，构建Skn1a真核表达载体，并转染Hela细胞，检测其表达情况。结果： RTPCR扩增得到约1 300 bp的cDNA 片段，将其克隆至pMD18T载体，DNA测序表明，得到片段长度为1 308 bp ,为转录因子Skn1a基因编码的cDNA，获得的Skn1a cDNA序列与已公布的Skn1a cDNA序列具有99.85%同源性, 有1个碱基的差异，但并未导致编码氨基酸的改变，继而构建了Skn1a真核表达载体pcDNA3/Skn1a，其转染Hela细胞后可正确表达。结论：转录因子Skn1a在宫颈癌组织中有一定量的表达，并在体外培养的Hela细胞中获得表达。 ??? 关键词： 宫颈癌；Skn1a；克隆；表达 ???? cDNA cloning and expressing of human Skn1a cDNA from ??? cervical carcinoma tissue ??? WANG Yanping,? MA Zhenghai,? YU Meng, et al ??? (Key Laboratory of Molecular Biology, College of Life Science and Technology, Xinjiang University, ??? Xinjiang Key Laboratory of Biological Resources and Genetic Engineering, Urumqi 830046, China) ??? Abstract: Objective: To clone cDNA of human transcription factor Skn1a (related to epidermal differentiation and HPV gene expression) gene from cervical carcinoma tissue and construct the eukaryotic expression vector of the Skn1a cDNA. Methods: The Skn1a cDNA fragment was amplified by RTPCR from total RNA that was extracted from cervical carcinoma tissue，the PCR product was cloned into pMD18T vector and the nucleotide sequence was analyzed, subsequently,? the eukaryotic expression vector of the Skn1a cDNA was constructed, and transfected Hela cells, the expression of Skn1a was examined. Results: A cDNA fragment about 1300 bp was amplified by RTPCR，the result of sequencing showed the fragment was 1 308 bp in length, containing a cDNA coding human Skn1a. The nucleotide sequence homology to the previously published sequence of human Skn1a was 99.85%, there was one conversion in nucleic acid sequences, but the conversion didnt induce the change of coding amino acid; the? eukaryotic expression vector of Skn1a can express correctly after transfecting Hela cells. Conclusions: The results suggested that Skn1a gene expresses at some level in th