Fine-mapping And Transcriptome Analysis Of Leafy Gene In Maize

The breeding of silage maize hybrid is an important part in the development of the forage maize industry.Due to its large leaf area and high biological yield,it is one of the elite germplasm resources for breeding high quality silage maize.In the present study,the segregating population including F2 and F3 individuals was used to finely map gene for leaf number above the primary ear.The segregating population was derived from the cross between leafy inbred line Y915 and normal inbred line Zheng58.These two inbre lines have nine and five leaves above the primary ear,respectively.In addition,RNA-Seq technique was used for transcriptome analysis in near isogenic lines(NIL)with different leaf number above the primary ear.The aim of the present study is to provide reference for revealing the genetic mechanism controlling leaf number above the primary ear of maize and breeding high-yielding and high-quality silage maize hybrids.The main results are as follows:(1)Preliminary mapping of genes for leaf number above the primary ear in maize.?BSA method was used to map the genes for leaf number above the primary ear as the following procedure.Two DNA pools were constructed using F2 individuals with less than five and more than twelve leaves above the primary ear,respsectivly.The F2 individuals were derived from the cross of Zheng58×Y915.Five hundred and ninty five SSR markers covering the whole genome were used to identify genotype of DNA mixed pool and parents.A total of seven co-segregation markers were found on chromosome 3,5 and 9,respectively.?A population including 190 F2 individuals derived from Y915×zheng58 was analyzed using maize 56K SNP array technology.High density genetic map was constructed.A total of seven QTL controlling leaf number above the primary ear were detected.These QTL located on chromosome 2,3,5,6,7 and 8.Comparing the results of the BSA method and F2 mapping population mapping results,it was found that a major QTL qLN3-2 was detected in both methods.This QTL had the highest LOD value,up to 34.93.It was located between SYN32259 and PZE-103182430,and explained 60.92%of the phenotypic variation.(2)Using map-based cloning method to fine mapping the qLN 3-2.Firstly,the recombinant individuals of qLN 3-2 were screened from 190 individuals based on the genotypic date collected using chip-genotyping method.Basded on the recombinant plants with less than six leaves and more than eleven leaves above the primar ear,the candidate gene was located between molecular marker PZE-103176493 and PZE-103181867.Secondly,using 184 F3 plants with four leaves above the primary ear,we located the candidate gene between marker 21-57 and InDel-58.Finally,using 1746 F3 plants with five leaves above the primary ear,we delimited the qLN3-2 to a chromosome region of 1.53 M between InDel-02 and Y250.(3)Using RNA-Seq technology to analyze the NILs.A total of 1203 differentially expressed genes were detected between the two NIL lines with different leaf number.The line with more leaves above the primary ear showed 611 up-regulated genes and 592 down regulated genes in the V4 period compared its counterpart.GO analysis showed that the significant enrichment GO Term in up-regulated genes was photosystem I,and the significant enrichment GO Term in down-regulated genes was transcription factor TFIID complex.Pathway enrichment analysis showed that the significant enrichment pathway in up-regulated genes was photosynthesis,and the significant enrichment pathway in down-regulated genes was DNA replication.The result of GO analysis and pathway enrichment analysis showed that leaf number above the primary ear has a greater impact on photosynthesis.