Post-GWAS in-silico and molecular validation of genic and Intergenic loci associated with cell-wall recalcitrance in Populus

A recently completed genome-wide association study utilizing >1,000 Populus trichocarpa genotypes and 30,160 SNPs yielded numerous loci highly associated with cell wall recalcitrance. In the current study, we used resequencing data from >50 Populus genotypes to characterize haplotypic structure surrounding these candidate loci based on linkage disequilibrium. We also identified regulatory elements surrounding intergenic SNPs and their putative downstream targets using sequence motif prediction sofware. On the molecular level, putative regulatory candidate genes were validated using multiple approaches including the examination of their tissue/organ expression patterns, the evaluation of  allelic effects on  expression of reporter genes for the lignin, S-monolignol, cellulose, and hemicellulose biosynthetic pathways using the Populus protoplast transient expression system, as well as the generation of stable transgenic lines. We will present current progress and implications of our findings on on-going efforts for feedstock improvement toward increased lignocellulosic biofuels production.