【摘要】 目的分离新的Ｂ细胞活化基因。方法采用差异显示反转录ＰＣＲ技术（ＤＤＲＴ－ＰＣＲ）对人扁桃体活化和静止Ｂ细胞ｍＲＮＡ的差异显示情况进行分析，对显示片段进行克隆并行Ｎｏｒｔｈｅｒｎ分析，对Ｎｏｒｔｈｅｒｎ杂交阳性的ｃＤＮＡ片段进行测序并比较同源性。结果差异显示分析共获得明显差异表达的ｃＤＮＡ片段６２条，选择主要表达于活化Ｂ细胞上的２０条ｃＤＮＡ片段克隆到ｐＧＥＭ－Ｔ载体上，并逐一对静止和活化Ｂ细胞ＲＮＡ进行Ｎｏｒｔｈｅｒｎ分析，其中６条ｃＤＮＡ片段在活化Ｂ细胞中呈Ｎｏｒｔｈｅｒｎ杂交阳性且与差异显示情况相符，对它们进行测序，然后通过国际互联网与ＧｅｎＢａｎｋ，ＥＭＢＬ和ＤＤＢＪ的ＤＮＡ数据库比较序列同源性，发现其中４个克隆与已发现的基因具明显同源性，一个克隆是新的，另一个克隆虽然与人Ｔ细胞分泌的趋化因子Ｉ－３０９同源性达９５％，但二者转录本不同。结论通过ＤＤＲＴ－ＰＣＲ分离到两个可能为新的Ｂ细胞活化基因的ｃＤＮＡ片段，这为进一步克隆新的Ｂ细胞活化基因奠定了基础。更多还原

【Abstract】 Objective To clone the novel B lymphocyte activation gene. Methods The differential display RT PCR (DDRT PCR) technique was applied to compare mRNA from human tonsil resting and activated B lymphocytes. The differential display cDNA fragments were recovered, reamplified and cloned into pGEM T vectors, then Northern blots were carried out to analyse those fragments. The positive cDNA fragments were sequenced. Results Sixty two differential display cDNA fragments were obtained. Twenty of the 62 cDNA fragments which were mainly or uniquely expressed on activated B lymphocytes were cloned into pGEM T vectors, among which 6 clones were confirmed expressible on activated B cells by Northern blot analysis and named EST XB11, XD11, YA1, ZD11, ZD12 and ZE1 respectively. The sequence analysis had been finished. Four clones (XB11, YA1, ZD11 and ZD12) had high homology with recorded genes and one (XD11) was novel by a computer search against GenBank, EMBL and DDBJ DNA databases. The clone (ZE1) had 95% homology with human T cell secreted chemokine I 309, but they had different transcript. Conclusions Two cDNA fragments (XD11, ZE1) are supposed to be potentially representive of novel B cell activation genes and this research lays a foundation for cloning novel B cell activation genes.更多还原