Differential Expression And Screening Of LncRNA And CircRNA In Testes Of Tibetan Pigs Brfore And After Sexual Maturity

Sexual maturity is an important sign of normal reproductive ability of Tibetan boars.In the process of testicular development and spermatogenesis in male animals,there are more cell synergies involved and the process is complex.Long non-coding RNAs and circular RNAs exist in humans,cows,mice,pigs and so on as non-coding RNAs.They are involved in the regulation process of testicular development and spermatogenesis.In the present research progress,although studies on the regulatory mechanisms of long non-coding RNAs and circular RNAs on the development of boars’ testis have been reported,such studies on the sexual maturity of Tibetan boars have not been reported.Therefore,testis at two different developmental stages of Tibetan pigs were studied before(2 months of age)and after(24 months of age).Using transcriptome sequencing and bioinformatics,differences in the expression of long non-coding RNAs and circular RNAs in testis were analyzed.To predict and analyze the potential biological functions of differentially expressed long non-coding RNAs and circular RNAs in testicular development and spermatogenesis,this study provides new clues to understand the role of long non-coding RNAs and circular RNAs in the regulation of spermatogenesis and testis development in Tibetan boars.The main research results are as follows:(1)Differential expression of long non-coding RNAs in testicular tissues of Tibetan pigs before and after sexual maturitySequencing was performed using Illumina sequencing platform,a total of63213 Mb Clean reads were obtained,6893 pig long non-coding RNAs,2775 newly predicted long non-coding RNAs,a total of 1875 long non-coding RNAs were screened,and their differential expression was significant(P< 0.05).Among them,660 long non-coding RNAs were significantly up-regulated and1215 long non-coding RNAs were significantly down-regulated.GO and KEGG enrichment analysis of differentially expressed long non-coding RNAs target genes showed that SPAG9,LOC100511430,SPACA1,STRBP,SPATA9,TDRP and TNP1 were related to male reproductive development of Tibetan pigs.Nine long non-coding RNAs that were differentially expressed before and after sexual maturity of Tibetan pigs were randomly screened for RT-q PCR.The results were basically consistent with the trend of transcripsomal sequencing analysis,indicating that the RNA-seq sequencing data were reliable.(2)Analysis of circular RNAs differentially expressed in testes of Tibetan pigs before and after sexual maturityA total of 16065 circular RNAs were identified.6317 circular RNAs were up-regulated and 9748 were down-regulated in the testes of Tibetan pigs before and after sexual maturity.By studying the interaction of circ RNAs-mi RNAs,the function of differentially expressed genes of circular RNAs bound to mi RNAs was identified by mi RNA host gene prediction.It is concluded that circular RNAs with different expression abundance exist in the testes of Tibetan pigs before and after sexual maturity.The host genes ICA1 L,STRBP,DPY19L2,TDRD5,FNDC3 A,MEIOB,TEX14,TDRKH,TEX15,KMT2 E and NSD2 may be involved in the biological processes of testicle development,spermatogenesis and sperm cell development.These circular RNAs can be used as biomarkers to predict fertility in boars.circular RNAs and mi RNAs bind more to the target sites of mi R-9858-5p、mi R-9813-5p and mi R-342,but no relevant studies have shown that they have certain promotion or inhibition effects on testicular development or spermatogenesis.The mi R-33b-3p family is involved in cell division,and related circular RNAs may act as mi RNA sponges to control testicular growth and spermatogenesis.In summary,this study analyzed the expression levels of long non-coding RNAs and circular RNAs in testicular tissues of Tibetan pigs before and after sexual maturity by RNA-seq technology.It was found that there were a large number of differentially expressed long non-coding RNAs and circular RNAs in testicular tissues before and after sexual maturity of Tibetan pigs,which were related to testicular development and spermatogenesis.To analyze,identify and verify the differences in the expression of these two non-coding RNAs in the male testicular tissue,The target genes that differentially expressed long non-coding RNAs SPAG9、LOC100511430、SPACA1、STRBP、SPATA9、TDRP、TNP1 and host genes that differentially expressed circular RNAs were ICA1L、STRBP、DPY19L2、TDRD5、FNDC3A in Tibetan pig testis tissue,MEIOB、TEX14 、 TDRKH 、 TEX15 、 KMT2 E and NSD2 are involved in testicle development and spermatogenesis.This study provides a theoretical basis for further revealing the regulatory role of non-coding RNAs in testicular development and spermatogenesis in Tibetan pigs.