| Wheat is one of the three major food crops in the world,and high yield and high quality are important breeding goals.QTL localization for grain weight-related traits(including grain length and width),identification of candidate genes,and the development of functional markers based on superior allelic variation can accelerate the process of molecular breeding.In this study,through the screening of the remaining heterozygous lines(RHL)of the progeny of the RIL population,a mapping population derived from RHL78,namely the HIF78(heterogeneous inbred families 78),was successfully constructed,and was carried out QTL analysis for grain weight-related traits.Combined with transcriptome sequencing,the candidate genes of major QTL regions were preliminarily verified.The main results were as follows:1.Important agronomic traits of the HIF78(354 lines)were identified,it is proved that there is a significant positive correlation between grain weight and grain shape,and the genes controlling the variation of traits are pleiotropic;863 pairs of SSR markers and 194 SNP markers were used for genotyping of HIF78 mapping population,and a total of 1 SSR marker and 7 SNP markers were screened for the construction of the genetic linkage map;Ici Mapping V4.1 software was used to construct genetic maps and perform QTL mapping for grain weight-related traits.2.A total of 8 QTLs for grain weight-related traits were detected on chromosome 3DS by the ICIM method,QGL.nwafu.3D.1,QGL.nwafu.3D.2,QGP.nwafu.3D,QPH.nwafu.3D,QGLW.nwafu.3D,QSL.nwafu.3D,QGA.nwafu.3D,and QTGW.nwafu.3D explained 32.11%,32.15%,54.86%,56.66%,40.73%,36.87%,36.27% and 18.89% of the phenotypic variation,respectively.These QTLs were located in almost the same genetic region,flanked by Marker6 and Marker10 and covering a physical interval of 3.8 Mb.It was obvious that this locus was a pleiotropic QTL that can affect grain shape,panicle shape and plant type and other traits,simultaneously.3.Through the transcriptome sequencing of RHL78 homozygous lines(RHL78＿L vs RHL78＿S).The results showed 28 high-confidence genes wereannotated in this QTL,and none of them exhibited significantly differential expression;but for sequence variation,there was only one reliable SNP allelic variation(C>G)in the ninth exon of gene Traes CS3D02G137200,which was named as Tasg-D2.Besides,the accuracy of the C>G allelic variation was further proved through the CDS verification of Tasg-D2 in parents Pindong34 and Warran.The SNP resulted in an amino acid substitution(Arg>Gly)in the TREE domainof Tasg-D2 which might be the key variation for the segregation of traits in the HIF78.Then,RT-q PCR was perfoemed for the tissue-specific expression of Tasg-D2 and its three homoeologues both in parents and homozygous lines(RHL78-L/S),the results showed that Tasg-D2 was most highly expressed in the parental young spikes;there was no significant difference in the expression of the three homoeologues of Tasg-D2 between the RHL78 pure lines,further validating the reliability of the transcriptome results.4.We successfully developed a d CAPS marker to confirm the genotyping effects of G>C allelic in the derived HIF78 mapping population,the results showed that the Tasg-D2-C accounted for 274/354,while the Tasg-D2-G accounted for 80/354,and the ratio of the two haplotypes was close to 3:1 and almost co-segregated with grain length to width ratio.Overall,compared with the lines with Tasg-D2-G,the lines with Tasg-D2-C had significantly higher1000-grain weight,grain length,grain width,grain length to width ratio,grain perimeter grain area,spike length and plant height,further illustrating the pleiotropic effects of this gene.Therefore,based on the obvious yield potential of this locus,the development of its functional markers could provide new choices for molecular marker-assisted selection in wheat,promoting the breeding and improvement of high-yielding wheat varieties in the future. |
