Mining Fiskeby III and Mandarin (Ottawa) Expression Profiles to Understand Iron Stress Tolerant Responses in Soybean.

The soybean (Glycine max L. merr) genotype Fiskeby III is highly resistant to a multitude of abiotic stresses, including iron deficiency, incurring only mild yield loss during stress conditions. Conversely, Mandarin (Ottawa) is highly susceptible to disease and suffers severe phenotypic damage and yield loss when exposed to abiotic stresses such as iron deficiency, a major challenge to soybean production in the northern Midwestern United States. Using RNA-seq, we characterize the transcriptional response to iron deficiency in both Fiskeby III and Mandarin (Ottawa) to better understand abiotic stress tolerance. Previous work by our group identified a quantitative trait locus (QTL) on chromosome 5 associated with Fiskeby III iron efficiency, indicating Fiskeby III utilizes iron deficiency stress mechanisms not previously characterized in soybean. We targeted 10 of the potential candidate genes in the Williams 82 genome sequence associated with the QTL using virus-induced gene silencing. Coupling virus-induced gene silencing with RNA-seq, we identified a single high priority candidate gene with a significant impact on iron deficiency response pathways. Characterization of the Fiskeby III responses to iron stress and the genes underlying the chromosome 5 QTL provides novel targets for improved abiotic stress tolerance in soybean.

A genome-wide analysis of the USDA Soybean Isoline Collection.

The USDA Soybean Isoline Collection has been an invaluable resource for the soybean genetics and breeding community. This collection, established in 1972, consists of 611 near-isogenic lines (NILs) carrying one or multiple genes conferring traits that had been determined to exhibit Mendelian inheritance. It has been used in multiple studies on the genetic basis, physiology, and agronomy of these qualitative traits. Here, we used publicly available genotype (SoySNP50K), phenotype, and pedigree data on this collection to characterize the isogenicity of the NILs and identify chromosomal positions of unmapped genes. A total of 368 NILs had at least 80% identity to their recurrent parent and, thus, were useful for what can be called introgression mapping. Both on-target and off-target introgressions were evaluated. The size of on-target introgressions into individual NILs ranged from 61 kb to 8.4 Mb, whereas off-target introgressions ranged from 2.6 kb to 54.8 Mb. The observed large off-target introgressions indicated that some NILs carry introgressions nearly the size of an entire chromosome. By applying introgression mapping to genes that had never been mapped, we identified the likely chromosomal positions of six such genes: ab, im, lo, Np, pc, and Rpm. The size of mapping intervals was large in some cases (10.28 Mb for im) but small in others (0.21 Mb for Np). The results reported herein will provide future researchers with a resource to help select informative NILs for future studies, and provide a starting point to further fine map, and ultimately clone and functionally characterize these six soybean genes.

Similar Seed Composition Phenotypes Are Observed From CRISPR-Generated In-Frame and Knockout Alleles of a Soybean KASI Ortholog.

The ß-ketoacyl-[acyl carrier protein] synthase 1 (KASI) gene has been shown in model plant systems to be critical for the conversion of sucrose to oil. A previous study characterized the morphological and seed composition phenotypes associated with a reciprocal chromosomal translocation that disrupted one of the KASI genes in soybean. The principle findings of this work included a wrinkled seed phenotype, an increase in seed sucrose, a decrease in seed oil, and a low frequency of transmission of the translocation. However, it remained unclear which, if any, of these phenotypes were directly caused by the loss of KASI gene function, as opposed to the chromosomal translocation or other associated factors. In this study, CRISPR/Cas9 mutagenesis was used to generate multiple knockout alleles for this gene, and also one in-frame allele. These soybean plants were evaluated for morphology, seed composition traits, and genetic transmission. Our results indicate that the CRISPR/Cas9 mutants exhibited the same phenotypes as the chromosomal translocation mutant, validating that the observed phenotypes are caused by the loss of gene function. Furthermore, the plants harboring homozygous in-frame mutations exhibited similar phenotypes compared to the plants harboring homozygous knockout mutations. This result indicates that the amino acids lost in the in-frame mutant are essential for proper gene function. In-frame edits for this gene may need to target less essential and/or evolutionarily conserved domains in order to generate novel seed composition phenotypes.

Identification and Fine-Mapping of a Soybean Quantitative Trait Locus on Chromosome 5 Conferring Tolerance to Iron Deficiency Chlorosis.

CORE IDEAS: 'Fiskeby III' harbors a combination of abiotic stress traits, including iron deficiency chlorosis (IDC) tolerance. An IDC quantitative trait locus on chromosome Gm05 was identified in genome-wide association studies and biparental populations. Fine-mapping resolved a 137-kb interval containing strong candidate genes. Iron deficiency chlorosis (IDC) is an important nutrient stress for soybean [Glycine max (L.) Merr.] grown in high-pH soils. Despite numerous agronomic attempts to alleviate IDC, genetic tolerance remains the most effective preventative measure against symptoms. In this study, two association mapping populations and a biparental mapping population were used for genetic mapping of IDC tolerance. Quantitative trait loci (QTLs) were identified on chromosomes Gm03, Gm05, and Gm06. Heterogenous inbred families were developed to fine-map the Gm05 QTL, which was uniquely supported in all three mapping populations. Fine-mapping resulted in a QTL with an interval size of 137 kb on the end of the short arm of Gm05, which produced up to a 1.5-point reduction in IDC severity on a 1 to 9 scale in near isogenic lines.

A comparison of pine and spruce in recovery from winter stress; changes in recovery kinetics, and the abundance and phosphorylation status of photosynthetic proteins during winter.

During winter evergreens maintain a sustained form of thermal energy dissipation that results in reduced photochemical efficiency measured using the chlorophyll fluorescence parameter Fv/Fm. Eastern white pine (Pinus strobus L.) and white spruce [Picea glauca (Moench) Voss] have been shown to differ in their rate of recovery of Fv/Fm from winter stress. The goal of this study was to monitor changes in photosynthetic protein abundance and phosphorylation status during winter recovery that accompany these functional changes. An additional goal was to determine whether light-dependent changes in light harvesting complex II (LHCII) phosphorylation occur during winter conditions. We used a combination of field measurements and recovery experiments to monitor chlorophyll fluorescence and photosynthetic protein content and phosphorylation status. We found that pine recovered three times more slowly than spruce, and that the kinetics of recovery in spruce included a rapid and slow component, while in pine there was only a rapid component to recovery. Both species retained relatively high amounts of the light harvesting protein Lhcb5 (CP26) and the PsbS protein during winter, suggesting a role for these proteins in sustained thermal dissipation. Both species maintained high phosphorylation of LHCII and the D1 protein in darkness during winter. Pine and spruce differed in the kinetics of the dephosphorylation of LHCII and D1 upon warming, suggesting the rate of dephosphorylation of LHCII and D1 may be important in the rapid component of recovery from winter stress. Finally, we demonstrated that light-dependent changes in LHII phosphorylation do not continue to occur on subzero winter days and that needles are maintained in a phosphorylation pattern consistent with the high light conditions to which those needles are exposed. Our results suggest a role for retained phosphorylation of both LHCII and D1 in maintenance of the photosynthetic machinery in a winter conformation that maximizes thermal energy dissipation.