关键词: 五指山猪, 肌纤维, 结构变异, 蛋白结构域, 相关性

Abstract:

The study aimed to explore the effect of structural variation (SVs) in MYH gene family on muscle growth for longissimus dorsi muscle (LDM) of Wuzhishan pigs, and to provide a theoretical basis for further understanding of the rule and molecular mechanism of muscle growth. The study subjects were Wuzhishan pigs, and Large White pigs were used as control group. Three Wuzhishan pigs (4-month-old sows, average weight was 14.2 kg) and three Large White pigs (4-month-old sows, average weight was 58.0 kg) were raised in the same environment for one week before slaughter, fed sufficient food and water, and all pigs were starved for 24 hours before slaughter. LDM of 6 pigs were collected, complete DNA was extracted, library was built and sequenced by PacBio platform, the sequencing data of each sample was about 40 Gb. The sequencing data was aligned to reference genome. Filter condition: 50 bp≤SVs≤10 kb; Total RNA was extracted from longissimus dorsi muscle of each pig, transcriptome sequencing was performed, and gene expression was calculated. The distribution position and number of SVs on chromosomes of Wuzhishan pig and Large White pigs were counted, the specific SVs of Wuzhishan pigs were screened, and the phylogenetic tree of MYH gene family was constructed, the structure domains were predicted, and the correlation between MYH gene expression and muscle fiber indicators was analyzed. The results showed as follows: 1) There were 7 specific MYH genes in Wuzhishan pigs, which were mainly distributed in chromosomes 3, 5, 6, 12 and 13, and chromosome 12 contained 3 MYH genes; 2) Phylogenetic tree showed that 7 MYH genes of Wuzhishan pigs differentiated into 2 branches, encoding non-muscle myosin and muscle myosin respectively, MYH9 and MYH11 genes, MYH1 and MYH4 genes had the highest homology; All 7 MYH genes contained the Motor domain, and 5 MYH genes contain the Myosin-N domain. The head domain of MYH gene (Myosin-N and Motor domain) was relatively conservative, and the middle and tail domain of MYH gene were different. The MYH1, MYH4 and MYH9 genes of Wuzhishan pigs contained SVs, while those of Large White pigs did not. Pearson coefficient statistics showed that the existence of SVs in MYH genes might be related to gene expression. The correlation between the expression levels of these three MYH genes and muscle fiber indexes was detected by both RNA-seq and Q-PCR. MYH1 gene expression was positively correlated with muscle fiber density and number, MYH4 gene expression was negatively correlated with muscle fiber density and number, MYH9 gene expression was negatively correlated with muscle fiber density and positively correlated with diameter. In summary, the largest number of MYH genes distributed on chromosome 12, the MYH gene family is mainly divided into two branches: non-muscle myosin and muscle myosin. The head domain is relatively conservative, and the difference between the middle and tail domain may be the main reason for the differentiation of MYH gene family. The expression levels of MYH1, MYH4 and MYH9 genes were significantly correlated with muscle fiber density, indicating that these genes might affect LDM growth through regulating the muscle fiber density. This study not only expands the content of molecular mechanism of muscle growth and development, but also provides reference for optimizing genetic breeding model related to structural variation.

Key words: Wuzhishan pigs, muscle fiber, structural variations, protein domains, correlations