秦川牛FADS基因家族克隆、生物信息学分析及组织表达研究

doi:10.16431/j.cnki.1671-7236.2024.03.002

摘要/Abstract

摘要： 【目的】试验旨在系统分析秦川牛脂肪酸去饱和酶(FADS)基因家族,阐明该家族成员在秦川牛不同组织及脂肪细胞不同分化时期的时空表达规律,为进一步探究其在牛脂肪沉积中的作用奠定基础。【方法】试验采集秦川牛不同组织样本,从肾周脂肪中分离前体脂肪细胞,通过PCR扩增FADS基因家族CDS区序列,利用生物信息学软件分析预测其序列特征。利用实时荧光定量PCR检测FADS基因家族成员在成年牛不同组织及肾周前体脂肪细胞分化过程中的时空表达谱。【结果】试验成功扩增出FADS基因家族3个成员(FADS1、FADS2和FADS3)CDS区,大小分别为1 425、1 335、1 332 bp。系统进化树结果显示,秦川牛FADS基因家族成员氨基酸序列与瘤牛相似性最高,与小鼠的亲缘关系较远。生物信息学预测结果显示,FADS基因家族均为较稳定的亲水性蛋白,二级结构主要由α-螺旋构成,三级结构与二级结构预测结果一致。FADS1和FADS2主要定位于内质网(44.4%、44.4%),FADS3主要定位细胞膜中(39.1%)。实时荧光定量PCR结果显示,FADS1、FADS3基因在肌肉组织表达量较高,FADS2基因在肝脏中表达量最高,显著高于其他组织(P＜0.05)。在脂肪细胞的成脂分化过程中,与其他时间点相比,FADS1和FADS2基因在第2天时表达量最高(P＜0.05),FADS3基因则在第2天时表达量最低(P＜0.05)。【结论】本研究成功克隆了秦川牛FADS基因家族成员FADS1、FADS2、FADS3,在瘤牛、山羊、绵羊等哺乳动物中高度保守。FADS基因家族均为较稳定的亲水性蛋白,FADS1、FADS3基因在肌肉组织中高表达,FADS2基因在肝脏组织中高表达。在脂肪细胞的成脂分化过程中,分化第2天时,FADS1、FADS2基因表达量最高,FADS3基因表达量最低。试验结果进一步揭示FADS基因家族通过调控脂肪细胞分化进而影响肉牛脂肪沉积作用机制研究奠定基础。

关键词: 秦川牛; FADS基因家族; 生物信息学; 表达

Abstract: 【Objective】 The purpose of this experiment was to systematically analyze the fatty acid desaturases (FADS) gene family in Qinchuan cattle,clarify the spatiotemporal expression patterns of these members in various tissues and different differentiation stages of preadipocytes,and lay a foundation for further exploring the function and regulatory roles of FADS gene family in bovine fat deposition.【Method】 Different tissue samples of Qinchuan cattle were collected,the preadipocytes were isolated from perirenal fat,and the CDS region sequences of FADS gene family were amplified by PCR,and the sequence characteristics were predicted by bioinformatics software.Real-time quantitative PCR was used to detect the temporal and spatial expression profiles of FADS gene family members in different tissues of adult cattle and during the differentiation of prerenal adipocytes.【Result】 The CDS region of 3 members of FADS gene family (FADS1,FADS2 and FADS3) was amplified successfully with the size of 1 425,1 335 and 1 332 bp,respectively.The phylogenetic tree results showed that the amino acid sequences of FADS gene family members of Qinchuan cattle had the highest similarity with Bos indicus,and were far related to Mus musculus.The results of bioinformatics prediction showed that the FADS gene family were stable hydrophilic proteins,and the secondary structure was mainly composed of alpha helix,and the tertiary structure was consistent with the predicted secondary structure.FADS1 and FADS2 were mainly located in the endoplasmic reticulum (44.4% and 44.4%),and FADS3 was mainly located in the cell membrane (39.1%).Real-time quantitative PCR results showed that the expression of FADS1 and FADS3 genes were higher in muscle tissue,and the expression of FADS2 gene was the highest in liver, and significantly higher than that in other tissues (P<0.05).With the adipogenic differentiation of adipocytes,compared to other differentiation time, the expression of FADS1 and FADS2 genes were the highest on days 2 (P<0.05),while the expression of FADS3 gene was the lowest on days 2 (P<0.05).【Conclusion】 FADS1, FADS2 and FADS3 of FADS gene family in Qinchuan cattle were successfully cloned, which were highly conserved Bos indicus, Capra hircus and Ovis aries and other mammals. FADS gene family were relatively stable hydrophilic proteins, FADS1 and FADS3 genes were highly expressed in muscle, and FADS2 gene was highly expressed in liver. In the process of adipogenic differentiation of adipocytes, FADS1 and FADS2 gene expressions were the highest and FADS3 gene expression was the lowest on days 2 of differentiation. The results laid a foundation for further research on the mechanism of FADS gene family influencing fat deposition in beef cattle by regulating adipocyte differentiation.

Key words: Qinchuan cattle; FADS gene family; bioinformatics; expression

中图分类号:

S823

尹思琦, 赵刚奎, 高海旭, 孙红红, 辛怡然, 昝林森, 赵春平. 秦川牛FADS基因家族克隆、生物信息学分析及组织表达研究[J]. 中国畜牧兽医, 2024, 51(3): 903-915.

YIN Siqi, ZHAO Gangkui, GAO Haixu, SUN Honghong, XIN Yiran, ZAN Linsen, ZHAO Chunping. Cloning, Bioinformatics Analysis and Tissue Expression of FADS Gene Family in Qinchuan Cattle[J]. China Animal Husbandry and Veterinary Medicine, 2024, 51(3): 903-915.

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