Jurkat细胞株TCRγ基因重排及多重引物扩增分析

Analysis of TCRγ gene rearrangement in Jurkat cell line and multiple primers amplification

目的分析Jurkat细胞株TCRγ基因重排特点,观察多重引物PCR扩增Jurkat细胞株TCRγ基因重排的效果。方法 TCRγ基因重排正向、反向引物配对组合,分别扩增Jurkat细胞株DNA,阳性PCR产物测序并比对分析;多重引物组合、降落式PCR扩增Jurkat细胞株TCRγ基因重排,比较多重引物与单对引物扩增效果。结果两组单对引物扩增产物电泳出现强阳性条带,测序并比对证实为TCRγ基因重排;与胚系基因比对发现,重排后TCRγ基因存在删除和增加的碱基序列;多重引物扩增产物中出现阳性条带的组合,其引物与单对引物组合一致。结论 Jurkat细胞中存在两种不同的TCRγ基因重排,重排序列体现了基因重排多样性。多重引物结合降落式PCR扩增TCRγ基因重排,扩增效果与单对引物一致。

Purpose To analyze the characteristics of TCRγ Gene rearrangement in Jurkat cell line,and observe the effect multiple primers in amplifying TCRγ Gene rearrangements of Jurkat cell line.Methods Sense and antisense primers for TCRγ gene rearrangements were paired and introduced to amplify DNA of Jurkat cell line,respectively.Positive PCR products were sequenced and the sequences were submitted to contrasting and analyzing.Multiple combinations of primers along with touch-down PCR were introduced to amplify TCRγ gene rearrangements of Jurkat cell line,comparing the effect of multiple primers PCR with that of the single paired primers.Results In amplification products of single pair of primers,two combinations showed strong bands in electrophoresis.Sequencing and contrasting analysis proved the products to be TCRγ gene rearrangements.Comparison with germinal gene sequence,the rearranged TCRγ gene implicated with deletion and addition of base sequence.For those presented positive bands in multiple primers amplification products,the primers applied were in accordance with that of single paired primers.Conclusion Two different TCRγ gene rearrangements exist in Jurkat cell line and the rearranged sequences reflect the diversity of TCRγ gene rearrangements.Multiple primers with touch-down PCR amplifying TCRγ gene rearrangement can achieve the same results as that of single paired primers.