猪肛门闭锁候选基因FN1的克隆及其编码区突变

Cloning and coding region mutation of FN1 as an anorectal malformation candidate gene in pigs

利用猪的SNP芯片筛选锁肛致病候选基因,在猪15号染色体候选区域筛选出纤连蛋白1(fibronectin 1,FN1)基因。分别用锁肛猪和正常猪cDNA池克隆得到FN1基因开放阅读框的7 437bp序列,鉴定3个选择性剪切外显子,并通过直接测序获得21个编码区单核苷酸多态位点(SNPs)。其中第22外显子的148A>G和第27外显子的50G>C引起氨基酸的改变(1 167Ile>Val,1 435Glu-Asp),且这2个SNPs在现有锁肛猪和正常猪样品中基因型频率和基因频率均差异显著。研究结果提示FN1基因的突变可能影响肛门直肠的正常发育过程,但还需要进一步的功能验证。

SNP(single nucleotide polymorphism)chips were applied to screen the candidate genes linked with anorectal malformation,and the Fibronectin1(FN1)gene on porcine chromosome 15 was captured as a candidate gene.By direct sequencing using the cDNA pools,we obtained the porcine FN1 mRNA sequence with a 7 437 bp ORF,encoding 2 478 aa.And 21 SNPs as well as three regions subject to alternative splicing were also identified.Of these SNPs,two(148A>G in exon 22,50G>C in exon27)were non-synonymous(1 167Ile>Val,1435Glu-Asp).PCR-RFLP analysis of these two SNPs suggested that the allele and genotype frequencies in the AA pigs and unaffected pigs were different(P<0.05).Therefore,FN1 mutations may affect the normal development of anorectal process,but the result need to be further verified.