China Animal Husbandry & Veterinary Medicine ›› 2021, Vol. 48 ›› Issue (8): 2713-2726.doi: 10.16431/j.cnki.1671-7236.2021.08.004

• Biotechnology • Previous Articles     Next Articles

Screening and Identification of the microRNAs Associated with Intramuscular Fat Deposition in Muchuan Black-bone Chicken

YU Shigang1, WANG Gang1, LIAO Juan1, XIE Wei2   

  1. 1. Engineering Research Center of Sichuan Province Higher School of Local Chicken Breeds Industrialization in Southern Sichuan, Leshan Normal University, Leshan 614000, China;
    2. Leshan Animal Disease Prevention and Control Center, Leshan 614000, China
  • Received:2020-12-01 Online:2021-08-20 Published:2021-08-19

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Abstract: This study was aimed to identify differentially expressed microRNA(miRNA) (DEM)in the breast muscle between high intramuscular fat (IMF) and low IMF content chickens, and explore the miRNA regulatory mechanisms involved in IMF deposition in chicken.120 Muchuan Black-bone chickens were employed in this study.High-throughput RNA sequencing (RNA-Seq) was performed to identify miRNA in the breast muscle between three high IMF (H group) and low IMF (L group) content chickens.The sequencing data was analyzed to identify DEMs.Six miRNAs were randomly selected for quantitative analysis using Real-time quantitative PCR.The results of IMF contents determination showed that the average IMF level of 120 chickens was 4.08 g per 100 g muscle, and there was extremely significant difference in IMF content between H and L groups (P<0.01).Results of 6 RNA-Seq libraries showed that more than 10 000 000 Clean reads were obtained for each library, and the Clean reads ratio was greater than 93%, more than 50% of small RNA (sRNA) ranged from 21 to 23 nt in length, and the 22 nt sRNA was the most abundant.The average GC content was 48.95%.These results indicated that the sequencing data were reliably produced in our study.The proportion of sRNA annotated as rRNA, tRNA, snRNA, snoRNA and repetitive sequence in the 6 RNA-Seq libraries was 2.74%, 1.85%, 6.21%, 1.58%, 2.82% and 2.81%, respectively.A total of 578 known miRNAs and 500 pre-miRNAs were identified from the 6 chicken sequence libraries, with 23 DEMs were identified between two groups, of which 16 were upregulated and 7 were downregulated.The prediction analysis of miRNA target genes showed that 23 miRNAs which were differentially expressed between two chicken groups targeted a total of 628 genes.GO analysis results showed that a total of 30 significantly enriched GO terms were identified, including 6 cellular components, 13 biological process and 11 molecular function.KEGG pathway analysis showed that the target genes were mainly involved with insulin signaling pathway, galactose metabolism, adipocytokine signaling pathway, glycosphingolipid biosynthesis, glycolysis and gtarch and sucrose metabolism.Real-time quantitative PCR results indicated that the expression of gga-miR-124a-3p and gga-let-7a-5p in H group was significantly higher than that in L group (P<0.01;P<0.05), the expression of gga-miR-19a-3p in H group was significantly lower than that in L group (P<0.01), the expression of gga-miR-6553-3p and gga-miR-128-3p in H group were significantly lower than that in L group (P<0.05), and 5 DEMs expression pattern were consistent with sequencing data.The results showed that gga-miR-124a-3p, gga-let-7a-5p, gga-miR-6553-3p, gga-miR-128-3p and gga-miR-19a-3p were associated with adipogenesis in animal, which might be important for IMF deposition in chicken and would provide a foundation for clarifying the molecular regulatory mechanisms of miRNA involved in IMF deposition in chicken.

Key words: chicken; intramuscular fat; miRNA; high throughput sequencing

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