为进一步了解和认识ATF4基因的功能, 揭示ATF4对猪脂肪代谢的影响, 寻找与肉质性状相关联的分子标记, 文章采用PCR方法扩增了ATF4基因部分序列, 通过序列比对发现在翻译起始密码子ATG下游159 bp处存在A159G转换, 通过PCR-AluⅠ-RFLP对大白猪、长白猪、梅山猪和通城猪进行酶切分型, 发现在大白猪和长白猪中均为AA基因型, 在梅山猪和通城猪中均为GG基因型。进一步对大白猪×梅山F2群体资源家系进行了酶切分型, 并分析该位点的多态性与生产性状的关系。结果表明, ATF4的多态性与臀部平均膘厚存在极显著相关(P<0.01), 与胸腰椎间膘厚、平均膘厚、眼肌高、眼肌面积存在显著相关(P<0.05)。采用Real-time PCR分析了ATF4基因在大白猪与梅山猪背最长肌不同发育阶段的表达模式。结果表明, ATF4基因在大白猪和梅山猪胚胎期65 d和出生后3 d中的表达水平相对都比较低, 且在两品种间无明显差异; 而在出生后60 d和120 d, ATF4基因在大白猪中与梅山猪均出现了上调表达, 并且在梅山猪中的相对表达水平要显著高于大白猪。研究结果为进一步深入研究猪ATF4基因在脂肪代谢中的分子机理奠定了基础。
In order to understand the function of gene ATF4 and identify new DNA markers involved in pig production traits, the cDNA fragment of porcine ATF4 was cloned and sequenced. Sequence comparison revealed an A159G substitu-tion downstream of the initiation codon (ATG). We then carried out PCR-AluⅠ-RFLP analysis in Large white, Landrace, Tongcheng and Meishan pigs, followed by association analysis in F2 “Large white ×Meishan” resource family. In all the individuals tested, Large White and Landrace pigs possessed the AA genotype, while Meishan and Tongcheng pigs possessed the GG genotype. Association analysis in F2 resource family showed that this site was highly associated with buttock fat thickness (BFT) (P<0.01) and had significant effect on thorax-waist fat thickness (TFT), average backfat thickness (ABT), loin eye height (LEH), and loin eye area (LEA)(P<0.05). Real-time PCR was used to analyze the expression patterns of porcine ATF4 gene in longissimus dorsi at different development stages of Large White and Meishan pigs. The results showed that the gene expression levels of ATF4 were low 65 days after conception and 3 days after birth, but no significant differences were observed in both breeds. Meanwhile, the expression levels of porcine ATF4 gene were up-regulated 60 days and 120 days after birth in both breeds and the expression level in Meishan pigs was obviously higher than that in Large White pigs. These data could lay the foundation for further study on the molecular mechanism of porcine ATF4 gene in lipid metabolism.
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