Immunoglobulin heavy chain switch region recombination within a retroviral vector in murine pre-B cells.
Immunoglobulin heavy chain switch region recombination within a retroviral vector in murine pre-B cells.
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小鼠前 B 细胞中逆转录病毒载体内的免疫球蛋白重链开关区重组。
DOI：
10.1002/j.1460-2075.1987.tb04793.x
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Marcu,KB
中科院分区：
文献类型：
--
作者：
Ott,DE;Alt,FW;Marcu,KB
我们以逆转录病毒载体ZN(SMU/S Gamma 2b)TK1为底物，检测了免疫球蛋白重链恒定区(CH)基因开关(S)在小鼠前B细胞中的重组活性。锌(SMU/S伽马2b)TK1除了含有位于小鼠SMU和S伽马2b之间的单纯疱疹病毒胸苷激酶(HTK)基因外，还含有一个新霉素(Neo)抗性基因。在G-418中选择稳定获得ZN(SMU/S Gamma 2b)TK1载体，并通过对药物溴脱氧尿苷(BUDR)的抗性选择开关区域在这些前体内重组。ZN(SMU/S Gamma 2b)TK1感染18-8tk和38B9tk-preB细胞系后，HTK基因失活的表观频率分别为5×10~(-5)和1×10~(-5)次/细胞/代，而G-418抗性的Ltd-K成纤维细胞则以4×10~(-8)的表观失活频率丧失HTK表型。Southern杂交分析表明，在所检测的所有前-B克隆中，开关重组导致了hTK基因的缺失，而在hTK-克隆中，hTK基因的缺失不是由S区域重组介导的。在24个Pre-B克隆中，有21个的重组涉及SMU和S伽马2b序列的简单重复部分。这些结果表明，插入到ZN(SMU/S伽马2b)TK1中的CH基因S区片段足以在S区串联重复序列中进行B细胞特异性重组/缺失。
We have employed a retroviral vector, ZN(Smu/S gamma 2b)tk1, as a substrate for detecting the presence of immunoglobulin heavy chain constant region (CH) gene switch (S) recombination activity in murine pre‐B cells. ZN(Smu/S gamma 2b)tk1 contains a neomycin (neo) resistance gene in addition to the herpes simplex virus thymidine kinase (Htk) gene which is positioned between murine Smu and S gamma 2b sequences. Stable acquisition of the ZN(Smu/S gamma 2b)tk1 vector was selected in G‐418 and switch region recombination within these proviruses was selected by resistance to the drug bromodeoxyuridine (BUdR). Fluctuation analyses of ZN(Smu/S gamma 2b)tk1 infected 18‐8tk‐ and 38B9tk‐ pre‐B lines revealed Htk gene inactivations with apparent frequencies of 5 X 10(‐5) and 1 X 10(‐5) events/cell/generation, respectively, while G‐418 resistant Ltk‐ fibroblasts lost the HTK phenotype at an apparent rate of 4 X 10(‐8). Southern blot analysis demonstrated that switch recombination caused the deletion of the Htk gene in all pre‐B clones examined while the loss of Htk in Ltk‐ clones was not mediated by S region recombination. In 21 out of 24 pre‐B clones, the recombinations involved the tandemly repetitive portions of the Smu and S gamma 2b sequences. These results demonstrate that the CH gene S region segments inserted into ZN(Smu/S gamma 2b)tk1 are sufficient for B‐cell‐specific recombination/deletion within the S region tandem repeats.