Differential Expression Of MiRNA And Related Regulatory Genes In Maize Inbred Lines Before And After Pollination

The ear traits are the key part of the corn's high yield.After pollination,the ear development speed is the fastest.Therefore,through the study of the differences in gene expression and development-related genes before and after pollination of maize ears,It is of great significance for the growth and development of the female ear during this period and its metabolic mechanism.microRNA?miRNA?is a type of small non-coding single-stranded RNA,about 22 bases in length.It plays an important role in plant development,metabolism and adaptation to different degrees of stress response?drought,low nitrogen,low phosphorus,waterlogging stress,heavy metal lead stress,sheath blight stress,etc.?.This study was based on HiSeq high-throughput sequencing technology combined with bioinformatics methods to study the changes in expression of RNA-seq and small RNA-seq before and after pollination of X178 and Ye478 materials.Differential expression multiples were used to screen mRNAs and miRNAs that were differentially expressed before and after pollination,and miRNAs were used to negatively regulate the target mRNAs.Correspondences between miRNA and target mRNAs were established,and GO and KEGG Pathways were analyzed for target and miRNAs.In this study,eight candidate genes were finally obtained.These genes may play an important role in the process of pollen development before and after pollination,providing some theoretical reference for the subsequent research on the development process before and after pollination.The results of the study are as follows:1.Through RNA-seq analysis,we obtained 11336 differentially expressed genes before and after X178 pollination,and 8721 differentially expressed genes before and after Ye478pollination?FDR?0.001,|log₂Ratio|?1?.2.For small RNA-seq analysis,small RNAs of 24 nt in X178 and Ye478 were predominant,with miRNAs concentrated at 21 nt or 22 nt.However,the number of miRNAs after pollination was significantly lower than that before pollination.It can be speculated that the miRNA regulates the target gene,and it is relatively active in the early stage of the ear development,and slightly reduced in the late pollination stage.The proportion of small RNA of each type in X178 and Ye478 was similar.The sequence of 21-24 nt before pollination of X178 accounted for 23.78%,21.33%,5.05%and 43.38%of the total sequence.After pollination,these four sequences accounted for 15.82%,11.17%,6.70%and 62.52%of the total sequence,respectively.The sequence of 21-24 nt before pollination of Ye478 accounted for 27.78%,17.17%,6.09%,and 43.72%of the total sequence,respectively.After the pollination,these four length sequences accounted for 21.36%,13.88%,7.07%,and 54.39%of the total sequence,respectively.3.By small RNA-seq analysis,there were 100 differentially expressed miRNAs before and after X178 pollination,belonging to 21 miRNA families;There were 98 miRNAs that were different before and after Ye478 pollination,belonging to 22 miRNA families?|fold change|?1,P<0.01?.81.0%of the miRNAs that were differentially expressed before and after X178 pollination had a miRNA MEFI?minimum folding free energy?value greater than 0.85.The MEFI values of 74.5%of the miRNAs that were differentially expressed before and after Ye478 pollination were greater than 0.85.It can be seen that the authenticity of most miRNAs is extremely high.In addition,in this study,it was found that of the 100 differentially expressed miRNAs in X178 and 63 of the 5'first base were U,accounting for 63.0%of the total.Among the 98 differentially expressed miRNAs of Ye478,51 of the 5'first base were U,accounting for 52.0%of the total.It can be seen that the first base of the miRNA mature sequence has a certain degree of bias toward base U,but the 10th and 11th base positions are more G and C.4.Through the integration analysis of RNA-seq and small RNA-seq,the corresponding relationship between miRNA and target mRNA was established.In X178 and Ye478,73 cross genes were obtained,of which 9 target genes did not find accurate gene names on the NCBI website.By performing GO enrichment analysis and KEGG Pathway analysis on the remaining 64 reliable target genes,8 reliable genes were finally obtained.These are GRMZM2G153233?ARF2?,GRMZM2G390641?ARF21?,GRMZM5G808366?ARF5?,GRMZ M2G081406?LOC103653810?,GRMZM2G058522?LOC103639134?,GRMZM2G110509,GRMZM2G153541 and GRMZM2G149768.Among them,GRMZM2G153233?ARF2?,GRMZM2G390641?ARF21?,GRMZM5G808366?ARF5?,GRMZM2G081406?LOC103653810?genes are regulated by zma-miR160a-5p,zma-miR160b-5p,and zma-miR160c-5p.The GRMZM2G058522?LOC103639134?gene is regulated by zma-miR398a-3p and zma-miR398b-3p.The GRMZM2G110509,GRMZM2G153541,and GRMZM2G149768?eEF1A?genes are regulated by miRNA169i-3p and miRNA169k-3p.These may be important candidate genes affecting the development of the ear before and after pollination.