DNA Methylation Analysis Of Photoperiod-B1in Wheat (Triticum Aestivum L.)

Photoperiod is a common phenomenon in plants and coordinated the internal circadianrhythms with external environment is very important for plant survival. Wheat (Triticumaestivum L.) is one of the world’s most important food crops which also affected byPhotoperiod. Photoperiod insensitivity (day neutral phenotype) enabled wheat adapt to abroad range of environments. Photoperiod-B1a is an important photoperiod insensitive allelewhich conferred day neutral phenotype in wheat. Previous studies suggested that copynumber variation (CNV) played a significant role in Ppd-B1allele’s formation. But theepigenetic modifications for this gene have not been reported. In this study, we furtherinvestigate the mechanism underlying photoperiod insensitivity allele Ppd-B1a from theepigenetic perspective—DNA methylation. The main progresses were as follows:(1) DNA methylation patterns in Ppd-B1promotor region showed rich polymorphisms.Using bisulfite genomic sequencing, we found DNA methylation patterns in Ppd-B1promotor region showed rich polymorphisms among a large germplasms. Two methylationhaplotypes (haplotype a&b) were classified according to the methylation polymorphisms ineach cytosine sites (single methylation polymorphisms, SMPs).(2) Methylation patterns varied between Ppd-B1methylation haplotypes.For the overall methylation patterns, haplotype b showed significantly highermethylation levels than haplotype a in both CG and CHG contexts. For each cytosine site, inCG and CHG motifs, haplotype b showed higher methylation levels in most positions (12/13and4/10). But for CHH, most cytosine site (29/40) showed no difference. In brief, themethylation levels were relative higher in haplotype b at both overall methylation level andeach cytosine site methylation level.(3) Ppd-B1methylation haplotypes associated with important agronomic traitsAssociation analysis was conducted between different methylation haplotypes andagronomic traits among105diverse wheat cultivars under three different environments in2011, China. And the results suggested that there were significant associations betweendifferent methylation haplotypes and agricultural traits. Compared with haplotype a,haplotype b associated with more favorable traits such as earlier heading time, reduced plantheight and higher1000-kernel weight. A linkage analysis was conducted between‘Fuzhuang30×Yanzhan1’ F7population methylation types and heading dates and the result showed that the heading dates of haplotype b was earlier than haplotype a. Both associationanalysis and linkage analysis suggested that the high methylation haplotype associated withearlier heading dates and conferred this gene photoperiod insensitive characteristics.(4) Ppd-B1methylation haplotypes underwent selection during wheat domestication.Different methylation haplotypes were differently distributed among wild species,landraces and modern cultivars. Low methylation haplotype predominate in ancient species.But in landraces and modern cultivars, high methylation haplotype became the leadinghaplotype which indicated that during wheat domestication high methylation patterns ofPpd-B1gene was selected.(5) There was no correlation between Ppd-B1methylation and copy number variationCorrelation analysis results showed that there were significant positive correlationsbetween different cytosine contexts methylation levels except CG and CHH. But there was nocorrelation between the relative copy number and methylation patterns in either cytosinemethylation context. So DNA methylation patterns and copy number variation were twoindependent factors affecting heading dates in wheat.(6) Positive correlation between Ppd-B1methylation and expressionQuantitative Real-time PCR results suggested that the expression level of Ppd-B1wasaffected by DNA methylation—high methylation pattern associated with high expression.Correlation analysis results showed that there were significant positive correlations betweenthe CG as well as the total methylation levels and the relative expression amounts.(7) Methylation difference is a unique characteristic for B genomeUsing bisulfite genomic sequencing, we detected the corresponding regions whichshowed different methylation status of Ppd-B1in A and D genomes. But the methylationpatterns showed no differences in Ppd-A1and Ppd-D1alleles which indicated that differentmethylation patterns might be a unique feature for Ppd-B1.