位置功能候选基因HMGA1、C6orf106和ENSSSCG00000023160与猪肢蹄结实度的关联性

doi:10.3864/j.issn.0578-1752.2016.20.016

摘要/Abstract

摘要： 【目的】建立一种通过内在的四肢骨关节评分来评估猪肢蹄结实度的方法，并计算关节评分与表观评估的肢蹄结实度之间的相关系数。此外，在F2、莱芜、二花脸、苏太和杜长大5个猪群中，研究三个位置功能候选基因HMGA1、C6orf106和ENSSSCG00000023160与猪肢蹄结实度之间的关联性。【方法】根据关节面裂痕的大小和深浅以及损伤的严重程度，对5块四肢骨的关节进行评分（1—5分）。如果关节面裂痕很大且很深，或损伤很严重，则评为1分；如果关节面没有裂痕和损伤，则评为5分。评分越高，说明关节越健康，肢蹄越结实。此外，基于笔者前期全基因组关联分析的结果，在猪7号染色体最强关联SNP两侧翼各0.2 Mb的区域内，筛选出HMGA1、C6orf106和ENSSSCG00000023160三个位置功能候选基因。在F2群体中，通过基因测序，搜寻这3个基因的多态位点，并根据多态位点在6个物种间的保守性，筛选出11个多态位点。利用Taqman探针，对3个HMGA1位点和3个C6orf106位点进行基因分型，而另一个基因的5个多态位点则通过基因型填补（genotype imputation）方法进行基因分型。最后，利用R软件GenABEL程序包，分析最小等位基因频率（MAF）大于0.05的多态位点与肢蹄结实度之间的关联性。【结果】在C6orf106和ENSSSCG00000023160基因中，分别鉴别到174和5个多态位点。关节评分之间呈显著的正相关，绝大部关节评分与蹄趾、肢蹄和步态评分之间无相关，而与肱二头肌长度和重量呈显著的负相关。公猪的肩胛骨关节评分极显著低于母猪的评分，但是臂骨肩关节和后肢跗关节评分显著高于母猪相应的关节评分。在F2群体中，3个候选基因均与肢蹄结实度关联，但是ENSSSCG00000023160的关联程度不如另外2个基因强，可以排除它为肢蹄结实度的因果基因。因此，该基因没有在其余的4个群体中进行检测。在二花脸群体中，HMGA1的g.2029C＞T和g.3155A＞G位点均与肢蹄结实度关联，另一个位点的MAF小于0.05。在其它3个群体中，HMGA1 3个位点的MAF都小于0.05。在莱芜群体中，仅C6orf106的g.6953T＞C位点的MAF大于0.05，该位点与肢蹄结实度显著关联。在苏太和杜长大群体中，C6orf106的g.2054T＞C和g.6953T＞C位点的MAF大于0.05，但它们与肢蹄结实度性状之间无显著关联。【结论】建立了一套利用四肢骨关节评分来评估肢蹄结实度的方法，该方法是对现有的表观评估方法的重要补充。因为蹄趾、肢蹄和步态评分与关节评分无相关，所以它们不能取代关节评分。关联分析结果排除了ENSSSCG00000023160是肢蹄结实度因果基因，但没有排除HMGA1和C6orf106的可能性，因此，有必要对这2个基因进行更深入的研究。

关键词: 猪, 肢蹄结实度, 关节评分, 关联分析, 基因型填补

Abstract: 【Objective】 The objective of this study is to develop a method to access the leg soundness through scoring the joint of limb bone and calculate the simple correlation coefficients among the scores for leg soundness in pigs. Furthermore, the association between three positional and functional candidate genes, namely HMGA1, C6orf106 and ENSSSCG0000023160, and leg soundness was also studied in F2, Laiwu, Erhualian, Sutai and DLY populations.【Method】The joint of five limb bones were scored according to the size and depth of rip on the joint surface and the worn-out degree of the joint. If the rip on the joint surface is very big and deep or the joint is seriously worn out, the joint is scored 1. On the other hand, if there is no rip on the joint surface and the joint doesn’t have any degree of worn-out, the joint is scored 5. Higher the joint score is, healthier the joint is and sounder the leg is. Based on the authors’ previous genome-wide association studies, three genes HMGA1, C6orf106 and ENSSSCG0000023160 were screened as positional and functional candidate genes to leg soundness in a 0.4 Mb region centered on the top SNP on SSC7. To search the polymorphic loci of the three genes in the F2 population, their DNA sequences were determined by a short-gun DNA sequence method. A total of 11 polymorphic loci were picked out according to their conservations among 6 species. The genotypes of 3 loci for HMGA1 and 3 loci for C6orf106 were determined using the Taqman method, and the genotypes of the other 5 loci for the third gene were inferred by genotype imputation just in the F2 population. At last, the GenABEL package of R was used to perform the association analysis between the loci with MAF＞0.05 and the traits.【Result】A total of 174 and 5 polymorphic loci were identified in C6orf106 and ENSSSCG00000023160 genes, respectively. Joint scores were positively correlated with each other and were negatively correlated with the length and weight of biceps brachii, but most of them had no correlation with toe, leg and gait scores. The male’s scapula joint score was significantly lower than the female’s, but arm shoulder joint score and focile hock joint score were significantly higher than the female’s corresponding joint score. In the F2 population, all of the three genes were associated with leg soundness, but ENSSSCG00000023160 was weaker than the other two genes, therefore it was excluded as a candidate gene to leg soundness and was not genotyped in the other 4 populations. In the Erhualian population, two loci g.2029C＞T and g.3155A＞G of HMGA1 were significantly associated with leg soundness, and the other SNP lacked the polymorphism. In the other 3 populations, all of the 3 SNPs of HMGA1 were deficiently polymorphic. Only the g.6953T＞C locus of C6orf106 had enough polymorphic in the Laiwu population, and it was associated with leg soundness. In the Sutai and DLY populations, only two loci g.2054T＞C and g.6953T＞C of C6orf106 were polymorphic, but none was associated with leg soundness.【Conclusion】A method of accessing the leg soundness has been proposed by scoring the joint of limb bones in pigs, and it is a crucial supplement method to access the leg soundness. Because toe, leg and gait scores are not correlation with the joint scores, they can’t replace the joint scores. The association analysis results excluded the ENSSSCG00000023160 gene as candidate gene to leg soundness, but both HMGA1 and C6orf106 genes were not excluded. Therefore, the two genes are worthy for further investigations.

Key words: pig, leg soundness, joint score, association analysis, genotype imputation

张徐非，候利娟，邱恒清，黄路生，郭源梅. 位置功能候选基因HMGA1、C6orf106和ENSSSCG00000023160与猪肢蹄结实度的关联性[J]. 中国农业科学, 2016, 49(20): 4030-4039.

ZHANG Xu-fei, HOU Li-juan, QIU Heng-qing, HUANG Lu-sheng, GUO Yuan-mei. An Association Study of Positional and Functional Candidate Genes HMGA1, C6orf106 and ENSSSCG00000023160 with Leg Soundness in Pigs[J]. Scientia Agricultura Sinica, 2016, 49(20): 4030-4039.

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