Additional file for paper Functional validation of gene IL6R and LEPR on reproductive traits in dairy cattle

Genetic selection of milk yield traits alters the energy distribution of high producing cows, resulting in gene-induced negative energy balance, and consequently, poor body condition scores and reduced reproductive performances. Here, we investigated two metabolic-syndrome pathway genes, <em>IL6R</em> (Interleukin 6 receptor) and <em>LEPR </em>(Leptin receptor)<em>,</em> for their effects on reproductive performance in dairy cows, by applying association analyses in 1,588 Chinese Holstein cows (at a population level) and gene expression analyses in granulosa cells isolated from eight cows (at a cell level). Of the six single nucleotide polymorphisms we examined (two SNPs for <em>IL6R</em> and four SNPs for <em>LEPR</em>), five were significantly associated with at least one reproductive traits, including female fetility traits covering both the abilitiy to to recycle after calving and the abilitiy to concive and keep pregnancy when inseminated properly, and calving traits. Notably, the identified variant SNP g.80143337A/C in <em>LEPR</em> is a missense variant. The role of <em>IL6R</em> and <em>LEPR</em> in cattle reproduction were further confirmed by observed differences in relative gene expression levels amongst granulosa cells with different developmental stages. Collectively, the functional validation of <em>IL6R</em> and <em>LEPR</em> performed in this study improved our understanding of cattle reproduction while provided important molecular markers for genomic selection of reproductive traits in high-yielding dairy cattle.

产奶性状的遗传选育改变了高产奶牛的能量分配格局，引发基因介导的能量负平衡，进而导致体况评分下降与繁殖性能降低。本研究针对两条代谢综合征通路基因——白细胞介素6受体基因（IL6R，Interleukin 6 receptor）与瘦素受体基因（LEPR，Leptin receptor）——展开研究，旨在解析其对奶牛繁殖性能的影响。研究通过群体层面的关联分析（基于1588头中国荷斯坦奶牛）与细胞层面的基因表达分析（基于8头奶牛的卵泡颗粒细胞）开展相关实验。本研究共检测6个单核苷酸多态性位点（SNPs，single nucleotide polymorphisms）：IL6R基因含2个位点，LEPR基因含4个位点。其中5个位点与至少1项繁殖性状存在显著关联，涵盖雌性繁殖性状（包括产后恢复能力、适宜输精条件下的受孕与妊娠维持能力）以及产犊性状。值得注意的是，LEPR基因中检测到的g.80143337A/C位点为错义突变位点。不同发育阶段卵泡颗粒细胞中IL6R与LEPR的相对基因表达量存在显著差异，进一步验证了这两个基因在奶牛繁殖过程中的调控作用。综上，本研究通过对IL6R与LEPR的功能验证，加深了我们对奶牛繁殖调控机制的认知，同时为高产奶牛繁殖性状的基因组选择提供了重要的分子标记。