一种新的人血管内皮生长因子基因的剪切形式

目的：检测人胚胎肺组织中新的血管内皮生长因子（VEGF）基因的可变剪切形式。方法：对胎儿肺组织的总RNA进行逆转录聚合酶链反应,克隆并测定扩增片段的序列。结果：用一对引物（引物5′端－21－7,3′端554－576）可扩增出3条片段,1条长度为正常的VEGF165（619bp) ,1条为正常的VEGF121（487bp),第3条为新的VEGF mRNA“异常”剪切片段,测序结果显示,此片段扩增长度为639bp,同样为VEGF165全长核苷酸序列,但在VEGF165的第三和第四外显子之间插入了长度为20bp的片段,经序列检索发现20bp的插入序列为滞留的第Ⅲ内含子终末序列,含内含子剪切信号,上述移码突变导致VEGF基因的阅读框架发生变化,在第四外显子中游出现终止密码子,结论：在一引产胎儿的肺组织中发现了新的VEGFmRNA可变剪切形式,其生物学意义有待进一步研究。

A novel alternative splicing isoform of vascular endothelial growth factor gene in human beings

OBJECTIVE: To investigate a new alternative splicing isoform of vascular endothelial growth factor (VEGF) gene in human bejings. METHODS: The total RNA of the lung tissue of a legally aborted 4-month-old human fetus was isolated and then amplified by RT-PCR. The amplified product was cloned into the pMD18-T plasmid and pcDNA3.1(-). Then sequence analysis was conducted. RESULTS: The electrophoresis of the RT-PCR product showed one short band of VEGF121 comprising of 487 bp and a long band with two fragments: one normal VEGF165 comprising of 619 bp and one fragment comprising of 639 bp which was the same VEGF165 nucleotide sequence with a 20 bp fragment inserted between exon 3 and exon 4. Sequence analysis showed that this 20 bp long nucleotide was inserted from the 3' end of the third intron that contained splicing signal, thus causing shift mutation in reading frame of VEGF gene and early appearance of the stop code UAG in the middle of exon 4. CONCLUSION: A new alternative splicing isoform of VEGF has been identified in the lung tissue of a legally aborted human fetus. Its biological significance remains to be further investigated.