DNA REPAIR AND SOMATIC MUTATION IN ANTIBODY VARIABLE GENES

DNA修复中的遗传缺陷与缺乏 超突变将是非常重要的，因为在体内没有 研究突变的模型。 已获得三个小鼠基因敲除系：PMS2和MSH2 错配修复小鼠和XPA核苷酸切除修复小鼠。 纯合子(-/-)基因敲除和产仔(+/+)小鼠正在接受免疫， DNA将从B细胞中提取。内源性VX和VH基因是 经聚合酶链式反应扩增、克隆、测序。的频率 基因敲除小鼠和对照小鼠的V基因和Constant基因的突变 比较一下。初步结果显示不同的突变模式 正常和错配修复缺陷小鼠。 用紫外光照射杂交瘤细胞。这些细胞是 然后在体外培养，并在不同的时间点收获。DNA是 制备，并与限制性内切酶和T4内切酶孵育， 裂解胸腺嘧啶二聚体。用凝胶分离裂解的DNA 用Southern印迹法鉴定了差异表达基因 分析。目前的结果表明，V基因具有较慢的 修复比二氢叶酸还原酶基因。

A correlation between a genetic defect in DNA repair and lack of hypermutation would be very significant, since there are no in vivo models to study mutation. Three knockout lines of mice have been obtained: PMS2 and MSH2 mismatch repair mice, and XPA nucleotide excision repair mice. Homozygous (-/-) knockout and littermate (+/+) mice are being immunized, and DNA will be prepared from B cells. Endogenous Vx and VH genes are being amplified by PCR, cloned, and sequenced. The frequency of mutation in the V and constant genes from knockout and control mice are compared. Preliminary results show different patterns of mutation in normal and mismatch repair deficient mice. Hybridoma cells were irradiated with ultraviolet light. The cells were then grown in vitro and harvested at different time points. DNA was prepared, and incubated with restriction enzymes and T4 endonuclease, which cleaves thymine dimers. Cleaved DNA was separated by gel electrophoresis, and different genes were identified by Southern blot analysis. Current results suggest that V genes have a slower rate of repair than the dihydrofolate reductase gene.