Study On Meat Quality Traits And MiRNA And MRNA Transcriptome Analysis Of The Longissimus Dorsi Of Changbai Mountain Semi-Wild Pig And Northeast Min Pig

Many referenes indicate that the difference of meat quality traits between the pig breeds is related to differential expression of genes.Therefore,to explore the differentially expressed genes in muscle between the pig breeds at the level of transcriptome is important to explore the molecular genetic mechanism of meat quality traits.In current study,the RNA-Seq sequencing technology was used to analyze the differentially expressed miRNA and differentially expressed genes in the longissimus dorsi of Changbai mountain semi-wild pig and Northeast Min pig,and the target genes of the differentially expressed miRNA were predicted.Through the correlation analysis of the differentially expressed miRNA and the differentially expressed genes,the functional genes and regulatory pathways which affected meat quality traits were screened out.The main results of this study were as follows:(1)The meat quality traits analysis of Changbai mountain semiwild pig and Northeast min pigThe carcass index analysis showed that the carcass weight,loin muscle area and backfat thickness index had no significant difference Changbai mountain semi-wild pig and Northeast min pig(P>0.05);The meat quality physical index analysis showed that the pH value of Changbai mountain semi-wild pig was significantly higher than that of Northeast min pig(P<0.05).The shear force of Changbai mountain semi-wild pig was significantly lower than that of Northeast min pig(P<0.05),and there was no significant difference between the aspects of color,drip loss and cooking loss of Changbai mountain semi-wild pig and Northeast min pig(P>0.05);The routine chemical analysis showed that the crude fat of Changbai mountain semi-wild pig was significantly higher than that of the Northeast min pig(P<0.05).The cholesterol content of Changbai mountain semi-wild pig was significantly lower than that of Northeast min pig(P<0.05);while the crude protein content,ash and moisture between Changbai mountain semi-wild pig and Northeast min pig showed no significant difference(P>0.05);The free amino acid analysis showed that of the 17 amino acids except for cystine,the other 16 kinds of amino acids in muscle had no significant difference between Changbai mountain semiwild pig and Northeast min pig,but there was an increasing trend about the values in Changbai mountain semi-wild pig;The fatty acid analysis showed that the content of unsaturated fatty acids(C14:1,C16:1,C18:1T,C18:1C,C20:1N9)in Changbai mountain semi-wild pig was significantly higher than that in Northeast min pig(P<0.05).Although there was no significant difference about other unsaturated fatty acids between the two breeds,there was still an increasing trend about C18:2C,C18:3N6,C20:2 C20:0,C20:3N6 etc.unsaturated fatty acids in Changbai mountain semi-wild pig.(2)The comparative analysis of muscle fiber characteristics and muscle tissue type between Changbai mountain semi-wild pig and Northeast min pigThe muscle histological characteristics analysis showed that the muscle fiber diameter and muscle fiber cross-sectional area of longissimus muscle and the biceps femoris of Changbai mountain semi-wild pig were significantly higher than that of Northeast min pig(P<0.05),and the muscle fiber density of longissimus dorsi and biceps femoris in Changbai mountain semi-wild pig was significantly lower than that in Northeast min pig(P<0.05);The muscle fiber type comparative analysis showed that the fast muscle proportion of Changbai mountain semi-wild pig was significantly higher than that of Northeast min pig(P<0.05).(3)The comparative analysis of the miRNA expressing profiles of longissimus muscle of Changbai mountain semi-wild pig and Northeast min pigThe miRNA library of six longissimus muscle of Changbai mountain semi-wild pig and Northeast min pig was successfully constructed in this study,and the identified 552 differentially expressed miRNA including 237 new prediction miRNA was analyzed by RNA-seq technique.There were 23 significantly differentially expressed miRNA in these differentially expressed miRNA,and compared to Northeast min pig,Changbai mountain semi-wild pig had 15 significantly down-regulated expressing miRNAs and 8 significantly up-regulated expressing miRNAs.The target gene prediction analysis revealed that these differential miRNAs had 3299 target genes;GO enrichment analysis found that these target genes were enriched to 5161 biological processes,657 cell components and 1254 molecular function;KEGG analysis showed that these predicted target genes of miRNA were involved in 292 signaling pathways.(4)The comparative analysis of mRNA transcriptome of longissimus dorsi of Changbai mountain semi-wild pig and Northeast min pigThe gene library of the longissimus dorsi of Changbai mountain semiwild pig and Northeast min pig was successfully constructed and analyzed by RNA-seq in this study.The result showed that there were total 17171 genes in longissimus dorsi of Changbai mountain semi-wild pig and Northeast min pig,including 619 genes expressed only in the Changbai mountain semi-wild pig,710 genes only in Northeast min pig;153 significantly differentially expressed genes were identified in the longissimus dorsi of Changbai mountain semi-wild pig and Northeast min pig,and 55 genes were up-regulated and 98 genes were down-regulated in Changbai mountain semi-wild pig;GO analysis showed that 153 differentially expressed genes were annotated to 33 GO entries,11 of which were annotated to the biological process,11 ones were annotated to the cellular components and 11 ones were annotated to the molecular function;KEGG analysis showed that these differentially expressed genes were enriched in 130 pathways,which were mainly related to the signaling pathway of fat metabolism.(5)qRT-PCR validation and correlation analysis of miRNA and mRNA transcriptome resultsIn this study,the RNA-Seq sequencing results of 3 miRNA and 3 genes randomly selected were verified by qRT-PCR,the results showed that the qRT-PCR results of miRNA and mRNA were consistent with RNA-Seq sequencing results,which showed that RNA-Seq sequencing results were reliable;The correlation analysis showed that the obtained 6 significantly differentially expressed miRNA corresponded to 10 significantly differentially expressed genes,of which 1 down-regulated expressing miRNA(ssc-miR-9785-5p)corresponded to 2 up-regulated differentially expressed genes(ENSSSCG00000004142,ENSSSCG00000026686),5 upregulated expressing miRNA(ssc-miR-1343,ssc-miR-150,ssc-miR-204,ssc-miR-493-5p.ssc-miR-504)corresponded to 8 down-regulated significantly differentially expressed genes(ENSSSCG00000005498,ENSSSCG00000030165,ENSSSCG00000010219,ENSSSCG00000005381,ENSSSCG00000006331,ENSSSCG00000005027,ENSSSCG00000000504,ENSSSCG00000009233);The GO analysis showed that these differential genes were involved in the process include lipid metabolism regulation,gene transcription regulation,metal ion binding regulation,endoplasmic reticulum components,skeletal muscle cell differentiation,thymus development and ureteric bud morphogenesis etc.;The KEGG analysis showed that these genes were involved in these pathways including adherens junctionpathway,glycerolipid metabolism pathway,glycerophospholipid metabolism pathway,hippopathway,legionellosis pathway,tight junction pathway,transcriptional misregulation in cancerpathway,protein digestion and absorbption pathway and metalloendopeptidases pathway etc.As stated previously,10 differentially expressed genes obtained in this study are closely related to meat quality difference of Changbai mountain semi-wild pig and Northeast min pig,which will be the main research content of in the further work.Among them,GPAT3 and PAPPA genes are closely related to meat quality traits and will be the target genes in the next step study.