Tissue Expression, SNPs Scanning of CNTF Gene and Its Association with Growth Traits in Hu Sheep (Ovis aries)
CHENG Jiang-Bo1, ZHANG De-Yin1, ZHANG Yu-Kun1, SONG Qi-Zhi1, LI Xiao-Long1, ZHAO Yuan1, XU Dan1, ZHANG Xiao-Xue1, LI Chong1, YUAN Lyu-Feng1, LI Fa-Di2,3, WANG Wei-Min1,*
1 College of Animal Science and Technology, Gansu Agricultural University, Lanzhou 730070, China; 2 College of Grassland Agricultural Science and Technology, Lanzhou University, Lanzhou 730000, China; 3 Gansu Mutton Sheep Breeding Biotechnology Engineering Laboratory, Minqin 733300, China
Abstract：Ciliary neurotrophic factor (CNTF) is a cytokine with multiple biological functions in vivo. It is an important candidate gene affecting muscle development. The aim of this study was to investigate the tissue expression pattern of CNTF gene in sheep (Ovis aries), scan its SNPs and analyze its association with growth traits in a large population. In this study, 1 399 Hu sheep with clear family tree and accurate phenotype records were raised in a specialized performance measurement house. Before morning feeding at 80, 100, 120, 140, 160 and 180 d, respectively, the weight of the sheep was weighed, and body length, body height, chest circumference and cannon circumference were measured at the same time. At the end of the assay period, whole blood samples of each sheep were collected by jugular vein for genomic DNA extraction. They were slaughtered immediately and tissue samples were taken. According to design the CNTF mRNA sequence specific primers, RNA from the collected tissue samples was extracted and reverse transcribed to cDNA, using qPCR technology to detect the CNTF gene expression in 10 tissues of Hu sheep. Genomic DNA was extracted from whole blood samples of the experimental population using genomic DNA extraction kit according to the instructions. Mixed DNA was used as the template to amplify the designed PCR primers. SNPs of the gene was scanned by mixed pool sequencing method, and the SPSS 23.0 software was used for correlation analysis with sheep growth traits. This results found a high positive association between growth traits, with a higher correlation between weight and body size indicators such as body length, body height, chest circumferences and cannon circumferences (P<0.01). The ovine CNTF gene was ubiquitously expressed, with expression observed in 10 tissues including heart, liver, spleen, lung, and kidney, with higher expression in liver and rumen. The sheep CNTF gene fragment was amplified and sequenced, a g.2576 C>G polymorphism locus was detected in intron 1 of the sheep CNTF gene. The SNP sites were genotyped by KASPar technique, and 3 genotypes were obtained, namely CC, CG and GG. In the experimental population, the genotype frequencies of CC, CG and GG genotypes were 0.13, 0.12 and 0.75, respectively, and the frequencies of alleles C and G were 0.19 and 0.81, respectively. The polymorphism information content of this locus was 0.26, which showed moderate polymorphism, and its heterzygosity (He), homozygosity (Ho) and effective number of alleles (Ne) were 0.31, 0.69 and 1.45, respectively. The results of trait association analysis showed that the polymorphism locus was significantly correlated with the growth traits such as body weight, body height, body length, chest circumference and cannon circumference of Hu sheep from 80 to 180 d (P<0.05). The body weight at 80, 100, 120, 140 and 160 d, body height at 80, 100 and 120 d and chest circumference at 160 d of individuals with GG genotype were significantly higher than those with CG genotype (P<0.05). The body height at 140, 160, 180 d, chest circumference at 80, 100, 140 and 180 d and cannon circumference at each stage were significantly higher in individuals with GG genotype than those with CG genotype and CC genotype (P<0.05). These results indicated that the CNTF gene g.2576 C>G locus could be used as a candidate molecular marker for breeding of growth traits in Hu sheep.
程江博, 张德印, 张煜坤, 宋其志, 李晓龙, 赵源, 徐丹, 张小雪, 李冲, 袁律峰, 李发弟, 王维民. 湖羊CNTF基因组织表达、SNPs扫描及其与生长性状关联分析[J]. 农业生物技术学报, 2022, 30(3): 506-516.
CHENG Jiang-Bo, ZHANG De-Yin, ZHANG Yu-Kun, SONG Qi-Zhi, LI Xiao-Long, ZHAO Yuan, XU Dan, ZHANG Xiao-Xue, LI Chong, YUAN Lyu-Feng, LI Fa-Di, WANG Wei-Min. Tissue Expression, SNPs Scanning of CNTF Gene and Its Association with Growth Traits in Hu Sheep (Ovis aries). 农业生物技术学报, 2022, 30(3): 506-516.
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