Du J, Zhu Y, Shanmugam A, Kenter A L
Department of Microbiology and Immunology (M/C 790), University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612-7344, USA.
Nucleic Acids Res. 1997 Aug 1;25(15):3066-73. doi: 10.1093/nar/25.15.3066.
The molecular mechanism of immunoglobulin switch recombination is poorly understood. Switch recombination occurs between pairs of switch regions located upstream of the constant heavy chain genes. Previously we showed that switch recombination breakpoints cluster to a defined subregion in the Sgamma3, Sgamma1 and Sgamma2b tandem repeats. We have developed a strategy for direct amplification of Smu/Sgamma3 composite fragments as well as Smu and Sgamma3 regions by PCR. This assay has been used to analyze the organization of Smu, Sgamma3 and a series of Smu/Sgamma3 recombination breakpoints from hybridomas and normal mitogen-activated splenic B cells. DNA sequence analysis of the switch fragments showed direct joining of Smu and Sgamma3 without deletions or duplications. Mutations were found in two switch junctions on both sides of the crossover point, suggesting that template switching is the most likely model for the mechanism of switch recombination. Statistical analysis of the positions of the recombination breakpoints in the Sgamma3 tandem repeat indicates the presence of two sub-clusters, suggesting non-random usage of DNA substrate in the recombination reaction.
免疫球蛋白类别转换重组的分子机制尚不清楚。类别转换重组发生在恒定重链基因上游的成对转换区域之间。此前我们表明，转换重组断点聚集在Sgamma3、Sgamma1和Sgamma2b串联重复序列中的一个特定子区域。我们开发了一种通过PCR直接扩增Smu/Sgamma3复合片段以及Smu和Sgamma3区域的策略。该检测方法已用于分析杂交瘤和正常丝裂原激活的脾B细胞中Smu、Sgamma3以及一系列Smu/Sgamma3重组断点的组织情况。转换片段的DNA序列分析显示Smu和Sgamma3直接连接，无缺失或重复。在交叉点两侧的两个转换连接点发现了突变，这表明模板转换是类别转换重组机制最可能的模型。对Sgamma3串联重复序列中重组断点位置的统计分析表明存在两个子簇，提示在重组反应中DNA底物的使用是非随机的。
