Genome-wide regulatory gene-derived SSRs reveal genetic differentiation and population structure in fiber flax genotypes.

We designed a set of 580 simple sequence repeat markers; 506 from transcription factor-coding genes, and 74 from long non-coding RNAs and designated them as regulatory gene-derived simple sequence repeat (ReG-SSR) markers. From this set, we could anchor 559 ReG-SSR markers on 15 flax chromosomes with an average marker distance of 0.56 Mb. Thirty-one polymorphic ReG-SSR primers, amplifying SSR loci length of at least 20 bp were chosen from 134 screened primers. This primer set was used to characterize a diversity panel of 93 flax accessions. The panel included 33 accessions from India, including released varieties, dual-purpose lines and landraces, and 60 fiber flax accessions from the global core collection. Thirty-one ReG-SSR markers generated 76 alleles, with an average of 2.5 alleles per primer and a mean allele frequency of 0.77. These markers recorded 0.32 average gene diversity, 0.26 polymorphism information content and 1.35% null alleles. All the 31 ReG-SSR loci were found selectively neutral and showed no evidence of population reduction. A model-based clustering analysis separated the flax accessions into two sub-populations-Indian and global, with some accessions showing admixtures. The distinct clustering pattern of the Indian accessions compared to the global accessions, conforms to the principal coordinate analysis, genetic dissimilarity-based unweighted neighbor-joining tree and analysis of molecular variance. Fourteen flax accessions with 99.3% allelic richness were found optimum to adopt in breeding programs. In summary, the genome-wide ReG-SSR markers will serve as a functional marker resource for genetic and phenotypic relationship studies, marker-assisted selections, and provide a basis for selection of accessions from the Indian and global gene pool in fiber flax breeding programs.

Identification and functional characterization of a flax UDP-glycosyltransferase glucosylating secoisolariciresinol (SECO) into secoisolariciresinol monoglucoside (SMG) and diglucoside (SDG).

BACKGROUND: Lignans are a class of diphenolic nonsteroidal phytoestrogens often found glycosylated in planta. Flax seeds are a rich source of secoisolariciresinol diglucoside (SDG) lignans. Glycosylation is a process by which a glycosyl group is covalently attached to an aglycone substrate and is catalyzed by uridine diphosphate glycosyltransferases (UGTs). Until now, very little information was available on UGT genes that may play a role in flax SDG biosynthesis. Here we report on the identification, structural and functional characterization of 5 putative UGTs potentially involved in secoisolariciresinol (SECO) glucosylation in flax. RESULTS: Five UGT genes belonging to the glycosyltransferases' family 1 (EC 2.4.x.y) were cloned and characterized. They fall under four UGT families corresponding to five sub-families referred to as UGT74S1, UGT74T1, UGT89B3, UGT94H1, UGT712B1 that all display the characteristic plant secondary product glycosyltransferase (PSPG) conserved motif. However, diversity was observed within this 44 amino acid sequence, especially in the two peptide sequences WAPQV and HCGWNS known to play a key role in the recognition and binding of diverse aglycone substrates and in the sugar donor specificity. In developing flax seeds, UGT74S1 and UGT94H1 showed a coordinated gene expression with that of pinoresinol-lariciresinol reductase (PLR) and their gene expression patterns correlated with SDG biosynthesis. Enzyme assays of the five heterologously expressed UGTs identified UGT74S1 as the only one using SECO as substrate, forming SECO monoglucoside (SMG) and then SDG in a sequential manner. CONCLUSION: We have cloned and characterized five flax UGTs and provided evidence that UGT74S1 uses SECO as substrate to form SDG in vitro. This study allowed us to propose a model for the missing step in SDG lignan biosynthesis.