CAJ | 학술논문

目的探索人胚胎肺组织中是否存在一种新的血管内皮生长因子(VEGF)基因可变剪切形式.方法对1例4月龄引产胎儿肺组织的总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 bp,3’端554～576 bp)可扩增出3条片段,一条为长度正常的VEGF165(619 bp)全长序列,另一条为长度正常的VEGF121(487bp)全长序列,第三条为新的VEGF mRNA异常剪切片段.测序检测显示,此扩增片段长度为639 bp,同样为VEGF165全长核苷酸序列,但在VEGF165的第3和第4外显子之间插入了长度为20bp的碱基片段.序列分析显示此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基因阅读框架发生改变,在第4外显子中游出现终止密码,故仅含-VEGF165外显子l～3及4部分序列,在蛋白水平上可能编码含97个氨基酸的新肽链.结论在一引产胎儿的肺组织中发现了新的VEGF mRNA可变剪切形式,其生物学意义有待进一步研究.
목적 탐색인배태폐조직중시부존재일충신적혈관내피생장인자(VEGF)기인가변전절형식.방법 대1례4월령인산태인폐조직적총RNA진행역전록취합매련반응、극륭병측정확증편단적서렬.결과 용일대인물(인물5’단-21～7 bp,3’단554～576 bp)가확증출3조편단,일조위장도정상적VEGF165(619 bp)전장서렬,령일조위장도정상적VEGF121(487bp)전장서렬,제삼조위신적VEGF mRNA이상전절편단.측서검측현시,차확증편단장도위639 bp,동양위VEGF165전장핵감산서렬,단재VEGF165적제3화제4외현자지간삽입료장도위20bp적감기편단.서렬분석현시차삽입서렬위체류적제Ⅲ내함자종말서렬,함내함자전절신호.상술이마돌변도치VEGF기인열독광가발생개변,재제4외현자중유출현종지밀마,고부함-VEGF165외현자l～3급4부분서렬,재단백수평상가능편마함97개안기산적신태련.결론 재일인산태인적폐조직중발현료신적VEGF mRNA가변전절형식,기생물학의의유대진일보연구.
Objective To characterize a new alternative splicing isoform of human vascular endothelial growth factor (VEGF)gene.Methods The total RNA was extracted from the lung tissue of a legally aborted 4-month-old fetus and amplified by RT-PCR.The amplified product was cloned into the plasmid pMD18-T and plasmid pcDNA3.1- for sequence analysis.Results Electrophoresis of the RT-PCR products displayed one short band for VEGF121 (487 bp) and a long band.The latter was dharacterized to contain two fragments:one was normal VEGF165 (619 bp),and the other (639 bp) had an identical nucleotide sequence to VEGF165 with a 20 bp fragment inserted between exons 3 and 4.Sequence analysis showed that this 20-bp nudeotide was inserted from the 3' end of the third intron containing a splicing signal thus causing shift mutation in the reading frame of VEGF gene and early appearance of the stop codon UAG in the middle of exon 4.Conclusion A new altemative splicing isoform of VEGF probably exists in the lung tissue of a legally aborted human fetus,and its biological significance remains to be further investigated.