Combined RNA-seq And ITRAQ Technology To Mine Genes Related To Cytoplasmic Male Sterility In Soybean

Heterosis utilization is an effective way to improve the yield of domestic soybean.However,the mechanism of cytoplasmic male sterility in soybean is still unclear,and the existing combinations cannot maximize heterosis.In this study,transcriptome and proteomics sequencing techniques were used to study the soybean sterile line 99629-24A(hereinafter referred to as 24A)and its restorer line 99629-24B(hereinafter referred to as 24B).Through the joint analysis of the two omics,the related genes affecting soybean fertility were explored,and the regulation mechanism of sterile genes was explored,which provided theoretical basis and data support for the study of the molecular mechanism of soybean cytoplasmic male sterility and the preparation of heterosis combinations.The main results are as follows :1.The physiological indexes of flowering tissues of soybean cytoplasmic male sterile line24 A and maintainer line 24 B at full flowering stage were determined,and it was speculated that the factors affecting the sterility of soybean 24 A were related to material metabolism disorder and abnormal enzyme activity.2.The RNA-Seq results of flowering tissues of soybean cytoplasmic male sterile line 24 A and maintainer line 24 B at full flowering stage showed that 1273 differentially expressed genes(DEGs)were identified.Among them,464 were up-regulated and 809 were down-regulated,indicating that the expression of most fertility-related genes in 24 A was inhibited rather than activated.Gene Ontology(GO)enrichment analysis of 1273 DEGs showed that a total of 285 DEGs were enriched into 473 terms,of which 64.2% were enriched into Biological Process(BP),14.5% were enriched into Cell Components(CC),and 21.1% were enriched into Molecular Function(MF).When P<0.05,a total of 119 terms were enriched,and 33.3%,30.9% and 35.7%terms were enriched in BP,CC and MF,respectively.The results of KEGG alignment showed that a total of 278 DEGs were enriched in 172 pathways,among which the starch and sucrose metabolic pathways(55 DEGs)and the mutual transformation of pentose and glucuronic acid(48 DEGs)were significantly enriched.In addition,the oxidative phosphorylation pathway and hormone signaling pathway were also enriched in more DEGs,and the interaction of these DEGs may affect the fertility of 24 A.The q RT-PCR results of 13 randomly selected DEGs showed that the RNA-Seq sequencing results were credible.3.The flowering tissues of soybean cytoplasmic male sterile line 24 A and maintainer line24 B at full flowering stage were analyzed by i TRAQ technology.A total of 475 differentially expressed proteins(DEPs)were found in the sterile line 24A(compared with the maintainer line24B),of which 204 DEPs were up-regulated and 271 DEPs were down-regulated.GO enrichment analysis showed that when P<0.05,a total of 291 proteins were enriched into 405 terms,of which 55.8% were enriched into BP,19.7% were enriched into CC,and 24.4% were enriched into MF.KEGG enrichment results showed that a total of 154 proteins were enriched into 79 pathways.When P<0.05,there were 13 significant enrichment pathways.4.The transcriptome and proteome results of soybean cytoplasmic male sterile line 24 A and maintainer line 24 B were analyzed,and 21 genes and proteins with consistent expression patterns were screened.GO analysis showed that in BP,the first five terms(primary metabolic process,subsequent organic matter metabolic process,metabolic process,macromolecular catabolic process,organic matter catabolic process)were all down-regulated.In CC,about 94%of the differentially expressed proteins in the first five terms(membrane,integral component of membrane,intrinsic component of membrane,membrane component,cell periphery)were down-regulated.In MF,about 95% of the differentially expressed proteins in the first five terms(catalytic activity,hydrolase activity,pectinesterase inhibitor activity,aspartate esterase activity,pectinesterase activity)were down-regulated.KEGG enrichment results showed that among the21 differentially expressed genes and proteins with the same expression trend,15 were enriched in four metabolic pathways: metabolic pathway,mutual transformation of pentose and glucuronic acid,glycosaminoglycan degradation,and ABC transporter.Among them,the unknown functional proteins in the glycosaminoglycan degradation pathway were up-regulated in the sterile line 24 A compared with the maintainer line 24 B,and the differential proteins in the other pathways were down-regulated.