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目的： 明确纤维化形成中调控Ⅰ型胶原基因高水平转录的启动子片段，逐段研究小鼠α２（Ⅰ）原胶原基因２ ｋｂ 的启动子序列。方法： 从小鼠α２（Ⅰ）原胶原基因转录起始点上游－ ２ ｋｂ～＋ ５４ ｂｐ 的片段中，取长度不等的片段作为启动子，与含氯霉素乙酰基转移酶（ＣＡＴ）报告基因的质粒组成６ 个重组体，脂质体转染法转染上述重组体至胶原产生细胞（ＮＩＨ３Ｔ３）和非胶原产生细胞（ＣＯＳ７）， ＣＡＴ－ＥＬＩＳＡ测定细胞ＣＡＴ表达水平以比较各重组体的启动子活性。结果： － ７８０～＋ ５４ ｂｐ 序列具有最高启动ＣＡＴ表达活性且有不完全细胞特异性，缺失近转录起始点５００ ｂｐ， 即－ ２～－ ０．５ ｋｂ 片段的启动子活性最低。结论： 近转录起始点５００ ｂｐ 的序列为小鼠α２（Ⅰ）原胶原基因启动激活所必需，－ ７８０～＋ ５４ ｂｐ 片段有高启动活性，并有望以此序列发现纤维化相关的特异ＤＮＡ 结合蛋白
OBJECTIVE: To determine the promoter region of high-level transcription of type Ⅰ collagen gene in fibrosis, and to study the promoter sequence of mouse α2 (Ⅰ) procollagen gene 2 kb one by one. Methods: The fragments of 2 kb to + 54 bp upstream of the transcription initiation site of α2 (Ⅰ) procollagen gene of mouse were taken as promoters, and the fragments containing the chloramphenicol acetyltransferase (CAT) reporter gene The recombinant plasmids were transfected into NIH3T3 cells and COS7 cells by Lipofectamine 2000 transfection method. CAT levels were measured by CAT-ELISA to compare the initiation of each recombinant Subactivity. Results: The sequence of - 780 bp + 54 bp had the highest activity of CAT expression and incomplete cell specificity. The deletion of transcription start site was 500 bp, ie, the promoter activity of - 2 - 0.5 kb fragment was the lowest. CONCLUSION: A sequence of 500 bp near the start of transcription is necessary for the activation of mouse α2 (Ⅰ) procollagen gene. - The 780 bp to + 54 bp fragment has high priming activity and is expected to find fibrotic related specific DNA binding protein