鼻咽癌细胞株SUNE1中EB病毒LMP1全基因克隆及部分序列的测定

DNA cloning and partial sequence analysis of EBV LMP1 gene isolated from human nasopharyngeal carcinoma cell line SUNE1

目的 研究广东地区鼻咽癌（ＮＰＣ） 细胞株ＳＵＮＥ１ 中ＥＢ病毒ＬＭＰ１ 基因的结构特征。方法 利用聚合酶链反应从ＳＵＮＥ１ 细胞基因组中扩增ＬＭＰ１ 全长ＤＮＡ，并克隆到ｐｃＤＮＡ３ 载体中，采用Ｓａｎｇｅｒ 双脱氧末端终止法测定ＳＵＮＥ１ＬＭＰ１ ３ 个外显子及２ 个内含子覆盖的序列，并与病毒原型Ｂ９５８ＬＭＰ１ 进行比较分析。结果 克隆到ｐｃＤＮＡ３ 中的ＳＵＮＥ１ＬＭＰ１ 全基因长度为２－６ ｋｂ，测序长度为１３１２ ｂｐ，与Ｂ９５８ＬＭＰ１ 比较，核苷酸与氨基酸的同源性分别为９８－５ ％ 和９６ ％ ，核苷酸及氨基酸变异主要在第３ 外显子，但无Ｃ端３４３～３５２ 密码子３０ 个碱基的缺失，第１ 外显子上有ＸｈｏⅠ酶切位点的丢失。结论 广东地区ＮＰＣ细胞株ＳＵＮＥ１ 中，病毒ＬＭＰ１ 基因与原型株之间尽管致瘤性存在差异，但仍具有较高的同源性。提示ＮＰＣ细胞中ＬＭＰ１ 的致瘤特征可能并非由于ＬＭＰ１

Objective To study the variation of EBV LMP1 encoding gene isolated from human nasopharyngeal carcinoma cell line SUNE1 in Guangdong.Methods The EBV LMP1 gene was amplified from the SUNE1 cell genome by PCR and then the recombinant vector was constructed by inserting the PCR fragment into pcDNA3. The encoding EBV LMP1 spanning from exon1 to exon3 in recombinant vector was sequenced by the Sequence Analyzer.Results The 1 312 bp encoding EBV LMP1 pieces in 2.6 kb PCR fragments were compared with the same EBV segments in B95 8 cell line. The data indicated that the rate of nucleotide sequence homology between the two fragments is 98.5% and the rate of amino acid is 96%. The restricted enzyme site of XhoⅠ in exon1 was deleted in SUNE1 but the 30bp deletion at the carboxyl terminus in most Chinese NPC LMP1 gene was not present. Conclusion Although the LMP1 gene derived from SUNE1 had greater tumorigenicity than that derived from B95 8 cell, the high homology rate of nucleotide and amino acid sequences between them indicated that it was not the result from the variation of certain nucleotide sites but the change in amino acid domain.