SNP/Indel Screening Analysis of Porcine Ovarian Granulosa Cells Treated with Follicular Fluid Exosomes
The aim of this study was to analyze the effect of follicular fluid exosomes on the genes of ovarian granulosa cells,to understand the regulation mechanism of follicular fluid exosomes during follicular development,and to provide a theoretical basis for the study of sow reproduc-tion.In this study,6-month-old binary sows with good health status and similar body weight were selected as experimental materials,and 150 ovaries were collected after slaughter,and follicular fluid exosomes and porcine ovarian granulosa cells(POGCs)were obtained using gradient centrifugation,and follicular fluid exosomes and porcine ovarian granulosa cells was co-cultured in vitro.Porcine ovarian granulosa cells(granulosa cell samples,GC,n=3)and porcine ovarian granulosa cells co-cultured with exosomes(granulosa-exosome co-culture samples,GCE,n=3)were sequenced by RNA-seq.The results showed that an average of 5.54X107 clean reads were obtained per sample in the GC and GCE groups,and the Q20 and Q30 quality scores were greater than 92％.A total of 1 310 979 SNP mutations and 104 498 InDel mutations were obtained in 6 samples in the GC and GCE groups,among which the number of homozygous SNP/Indel mutations was 170 426 and the number of heterozygous SNP/Indel mutations was 1 245 052,and the number of mutant heterozygotes was significantly higher than the number of homozygotes and with a high incidence of SNPs.In addition,among the mutation types,there were 973 003 transformations and 339 974 transversion,and the number of transformations was significantly higher than the number of transversion.After gene annotation,the mutations mainly occurred in the 3'UTR,5'UTR,and intron regions of the genes,followed by exons and intergenic regions.In addition,after comparison with the differential genes,the 1 583 candidate genes were screened,and it was found by GO and KEGG functional enrichment that the candidate genes were mainly related to cell cycle and cell proliferation/apoptosis processes.In addition,a total of 14 key candidate genes related to cell cycle and proliferation/apoptosis pathways were identified,of which 11 genes had interaction relationships.These SNP/Indel information obtained in this study can provide a scientific basis for subsequent studies on the regulation of exosomes on sow reproduction.
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