Project description:The soybean (Glycine max) seed coat has distinctive, genetically programmed patterns of pigmentation and the recessive k1 mutation can epistatically overcome the dominant I and i-i alleles, which inhibit seed color by producing small interfering RNAs (siRNAs) targeting chalcone synthase (CHS) mRNAs. Small RNA sequencing of dissected regions of immature seed coats demonstrated that CHS siRNA levels cause the patterns produced by the i-i and i-k alleles of the I locus, which restrict pigment to the hilum or saddle region of the seed coat, respectively. To identify the K1 locus, we compared RNA-Seq data from dissected regions of two Clark isolines having similar saddle phenotypes mediated by CHS siRNAs but different genotypes (homozygous i-k K1 versus homozygous i-i k1). By examining differentially expressed genes, mapping information, and genome resequencing, we identified a 129-bp deletion in Glyma.11G190900 encoding Argonaute5 (AGO5), a member of the Argonaute family. Amplicon sequencing of several independent saddle pattern mutants from different genetic backgrounds revealed independent lesions affecting AGO5, thus establishing Glyma.11G190900 as the K1 locus. Non-functional AGO5 from k1 alleles leads to altered distributions of CHS siRNAs, thus explaining how the k1 mutation reverses the phenotype of the seed coat regions from yellow to pigmented, even in the presence of the normally dominant I or i-i alleles.
Project description:Two Near Isogenic soybean (Glycine max) lines were grown in hydroponic conditions with either 50uM ferric nitrate or 100uM ferric nitrate. After 10 days, half the plants were harvested (total root tissue). At 12 days after planting, iron was added to plants grown in low iron conditions bringing them up to sufficient iron growth conditions. Root tissue was harvested for the remaining plants at 14 days after planting. Gene expression analysis from root tissue of two Near Isogenic Lines (NILs), Clark (PI548553) and IsoClark (PI547430), grown in iron stress or iron stress recovered conditions.