RHD mRNA替代剪接区域研究

目的2007年国内报道一例弱D型个体存在第4—9外显子选择性剪接的转录子，我们探讨正常Rh（D）阳性个体的RHD基因mRNA的选择性剪接区域。方法随机选择3名Rh（D）阳性个体，从新鲜全血中提取总RNA，通过特异性引物，采用“一步法”逆转录-PCR（1iT—PCR），扩增RHDmRNA第1～7外显子区域，以及第6-10外显子区域，然后cDNA琼脂糖凝胶电泳和成像分析。结果未发现第1～7外显子区域存在mRNA的选择性剪接条带，仅存在由特异性引物所扩增的第1—7外显子全长的序列条带；而第6～10外显子区域观察到5种替代剪切条带，序列分析显示分别为无缺失片段，以及完整缺失第7、第9、第7和9、第7—9外显子5种RHD转录子。结论正常Rh（D）抗原阳性个体的RHD基因mRNA的选择性剪接仅存在于第7～9外显子区域。

A Study on the Alternative Splicing Regions of RHD mRNA

Objective It was reported in 2007 that the alternative splicing regions between ex- ons 4-9 exists in a weak D type individual. Here we sought to investigate the regions in RHD gene mRNA in normal Rh-positive individuals. Methods Total mRNA from whole blood was isolated from three normal Rh （D） -positive individuals. The region of RHD mRNA from exon 1 to 7 and the region from exon 6 to 10 were amplified through a one-step RT-PCR method. Then cDNA were sepa- rated through eleetrophoresis. Results A single cDNA band for the full length region of exon 1-7 was observed in all three D-positive individuals. However, 5 alternative bands were detected for the region of exon 6-10, which were fragments representative for the sequence with no deletion, and the sequences with full deletion of exon 7, exon 9, exons 7 and 9, exons 7-9. Conclusion Our data suggested that the RHD mRNA alternative splicing regions in normal D-positive individual occur in exons 7-9.