青稞早抽穗主效QTL cqHD2H-2的遗传定位及候选基因分析

Genetic mapping and candidate gene analysis of the major QTL cqHD2H-2 for early heading in barley(Hordeum vulgare L.)

抽穗期是决定青稞品种的种植区域范围和季节适应性的重要农艺性状,也是青稞成熟早晚的关键标志。在前期实验中,青稞早抽穗主效QTL cqHD2H-2被定位于青稞2H染色体上约84 Mb的区间内,为进一步验证该位点的有效稳定性,本研究对cqHD2H-2进行定位验证及候选基因分析。利用早抽穗青稞品种DZZ与晚抽穗青稞品种KL10构建F5群体,在cqHD2H-2区段内开发InDel标记,共筛选到3个与目标基因紧密连锁的分子标记,扫描F5群体中的25株极端早抽穗和25株极端晚抽穗单株,获得1个侧翼共显性标记,将cqHD2H-2进一步限定在标记PA22和Va07之间约40 Mb的区间内。该区段通过与大麦及水稻同源共线性比对,筛选到水稻抽穗期基因DTH8在大麦中的同源基因HORVU2Hr1G087460(HvNF-YB3)。比较HvNF-YB3基因全长,发现Hv2H.NF-YB3(DZZ)与Hv2H.nf-yb3(KL10)编码区由1个外显子组成,编码区为750 bp,启动子区为2116 bp,Hv2H.NF-YB3与Hv2H.nf-yb3启动子区存在3个SNPs。Hv2H.NF-YB3与Hv2H.nf-yb3的表达分析表明,Hv2H.NF-YB3与Hv2H.nf-yb3在转录水平上存在显著差异,Hv2H.NF-YB3与Hv2H.nf-yb3在茎、芒和颖壳组织中均有表达,但相较于Hv2H.NF-YB3,Hv2H.nf-yb3在各组织中的表达量均显著下降,且二者随着生育期的推进,表达量均依次降低。因此,Hv2H.NF-YB3可能在青稞抽穗时间调控中发挥作用。本研究为青稞早抽穗分子标记辅助选择育种体系的建立及目标基因的图位克隆奠定了基础。

Heading date is closely linked with agronomic performance,which is responsible for the regional and seasonal adaptation of barley varieties,and it is also a key indicator of the early or later maturity in barley.Previous studies indicated that the major QTL cqHD2H-2 for early heading was located in an 84 Mb interval on chromosome 2H of barley.To further verify the effective stability of the cqHD2H-2,in this study,we validated this locus and analyzed candidate genes.New InDel markers were developed based on the primary mapping region of cqHD2H-2 through an F5 population constructed from the cross of DZZ(early heading)and KL10(late heading).A total of three markers tightly linked to the target gene were obtained,and one flank marker was confirmed as co-dominant marker by scanning 25 extreme early heading and 25 extreme late heading individuals of F5 population,which mapped the locus cqHD2H-2 located in a 40 Mb region between PA22 and Va07 on the chromosome 2H.The homologous gene HORVU2Hr1G087460(HvNF-YB3)of rice heading date gene DTH8 in barley was acquired by homologous collinearity comparison of the region between barley and rice.Compared with the full length of HvNF-YB3 gene,the encoding region of Hv2H.NF-YB3(DZZ)and Hv2H.nf-yb3(KL10)was composed of an exon.The encoding region was 750 bp,and the promoter region was 2116 bp.There were three SNPs detected between the promoter region of Hv2H.NF-YB3 and Hv2H.nf-yb3.The relative expression levels of Hv2H.NF-YB3 and Hv2H.nf-yb3 showed that there were significant differences in the transcriptional level.Hv2H.NF-YB3 and Hv2H.nf-yb3 expressed in stem,awn,and glume,compared with Hv2H.NF-YB3,the relative expression levels of Hv2H.nf-yb3 significantly decreased in the three tissues.The relative expression levels gradually decreased with the development of the plants.In conclusion,Hv2H.NF-YB3 may play a role in the regulation of heading time in barley.The results were useful for developing early heading lines by marker-assisted selection(MAS)and laid a foundation for fine mapping and subsequent map-based cloning of cqHD2H-2.