Genotyping-by-Sequencing and QTL Mapping of Biomass Yield in Two Switchgrass F1 Populations (Lowland x Coastal and Coastal x Upland).

The prevalence of genetic diversity in switchgrass germplasm can be exploited to capture favorable alleles that increase its range of adaptation and biomass yield. The objectives of the study were to analyze the extent of polymorphism and patterns of segregation distortion in two F1 populations and use the linkage maps to locate QTL for biomass yield. We conducted genotyping-by-sequencing on two populations derived from crosses between the allotetraploid lowland genotype AP13 (a selection from "Alamo") and coastal genotype B6 (a selection from PI 422001) with 285 progeny (AB population) and between B6 and the allotetraploid upland VS16 (a selection from "Summer") with 227 progeny (BV population). As predictable from the Euclidean distance between the parents, a higher number of raw variants was discovered in the coastal × upland BV cross (6 M) compared to the lowland × coastal AB cross (2.5 M). The final number of mapped markers was 3,107 on the BV map and 2,410 on the AB map. More segregation distortion of alleles was seen in the AB population, with 75% distorted loci compared to 11% distorted loci in the BV population. The distortion in the AB population was seen across all chromosomes in both the AP13 and B6 maps and likely resulted from zygotic or post-zygotic selection for increased levels of heterozygosity. Our results suggest lower genetic compatibility between the lowland AP13 and the coastal B6 ecotype than between B6 and the upland ecotype VS16. Four biomass QTLs were mapped in the AB population (LG 2N, 6K, 6N, and 8N) and six QTLs in the BV population [LG 1N (2), 8N (2), 9K, and 9N]. The QTL, with the largest and most consistent effect across years, explaining between 8.4 and 11.5% of the variation, was identified on 6N in the AP13 map. The cumulative effect of all the QTLs explained a sizeable portion of the phenotypic variation in both AB and BV populations and the markers associated with them may potentially be used for the marker-assisted improvement of biomass yield. Since switchgrass improvement is based on increasing favorable allele frequencies through recurrent selection, the transmission bias within individuals and loci needs to be considered as this may affect the genetic gain if the favorable alleles are distorted.

Mapping freezing tolerance QTL in alfalfa: based on indoor phenotyping.

BACKGROUND: Winter freezing temperature impacts alfalfa (Medicago sativa L.) persistence and seasonal yield and can lead to the death of the plant. Understanding the genetic mechanisms of alfalfa freezing tolerance (FT) using high-throughput phenotyping and genotyping is crucial to select suitable germplasm and develop winter-hardy cultivars. Several clones of an alfalfa F1 mapping population (3010 x CW 1010) were tested for FT using a cold chamber. The population was genotyped with SNP markers identified using genotyping-by-sequencing (GBS) and the quantitative trait loci (QTL) associated with FT were mapped on the parent-specific linkage maps. The ultimate goal is to develop non-dormant and winter-hardy alfalfa cultivars that can produce extended growth in the areas where winters are often mild. RESULTS: Alfalfa FT screening method optimized in this experiment comprises three major steps: clone preparation, acclimation, and freezing test. Twenty clones of each genotype were tested, where 10 samples were treated with freezing temperature, and 10 were used as controls. A moderate positive correlation (r ~ 0.36, P < 0.01) was observed between indoor FT and field-based winter hardiness (WH), suggesting that the indoor FT test is a useful indirect selection method for winter hardiness of alfalfa germplasm. We detected a total of 20 QTL associated with four traits; nine for visual rating-based FT, five for percentage survival (PS), four for treated to control regrowth ratio (RR), and two for treated to control biomass ratio (BR). Some QTL positions overlapped with WH QTL reported previously, suggesting a genetic relationship between FT and WH. Some favorable QTL from the winter-hardy parent (3010) were from the potential genic region for a cold tolerance gene CBF. The BLAST alignment of a CBF sequence of M. truncatula, a close relative of alfalfa, against the alfalfa reference showed that the gene's ortholog resides around 75 Mb on chromosome 6. CONCLUSIONS: The indoor freezing tolerance selection method reported is useful for alfalfa breeders to accelerate breeding cycles through indirect selection. The QTL and associated markers add to the genomic resources for the research community and can be used in marker-assisted selection (MAS) for alfalfa cold tolerance improvement.

Correction to: QTL mapping of flowering time and biomass yield in tetraploid alfalfa (Medicago sativa L.).

In the article [1], in 'Methods' section and 'G x E and heritability' subsection, there is an error in the formula of heritability (H2).

QTL mapping of flowering time and biomass yield in tetraploid alfalfa (Medicago sativa L.).

BACKGROUND: The genetic and genomic basis of flowering time and biomass yield in alfalfa (Medicago sativa L.) remains poorly understood mainly due to the autopolyploid nature of the species and the lack of adequate genomic resources. We constructed linkage maps using genotyping-by-sequencing (GBS) based single dose allele (SDA) SNP and mapped alfalfa timing of flowering (TOF), spring yield (SY), and cumulative summer biomass (CSB) in a pseudo-testcross F1 population derived from a fall dormant (3010) and a non-dormant (CW 1010) cultivars. We analyzed the quantitative trait loci (QTL) to identify conserved genomic regions and detected molecular markers and potential candidate genes associated with the traits to improve alfalfa and provide genomic resources for the future studies. RESULTS: This study showed that both fall dormant and non-dormant alfalfa cultivars harbored QTL for early and late flowering, suggesting that flowering time in alfalfa is not an indicator of its fall dormancy (FD) levels. A weak phenotypic correlation between the flowering time and fall dormancy (FD) in F1 and checks also corroborated that alfalfa FD and TOF are not the predictors of one another. The relationship between flowering time and alfalfa biomass yield was not strong, but the non-dormant had relatively more SY than dormant. Therefore, selecting superior alfalfa cultivars that are non-dormant, winter-hardy, and early flowering would allow for an early spring harvest with enhanced biomass. In this study, we found 25 QTL for TOF, 17 for SY and six QTL for CSB. Three TOF related QTL were stable and four TOF QTL were detected in the corresponding genomic locations of the flowering QTL of M. truncatula, an indication of possible evolutionarily conserved regions. The potential candidate genes for the SNP sequences of QTL regions were identified for all three traits and these genes would be potential targets for further molecular studies. CONCLUSIONS: This research showed that variation in alfalfa flowering time after spring green up has no association with dormancy levels. Here we reported QTL, markers, and potential candidate genes associated with spring flowering time and biomass yield of alfalfa, which constitute valuable genomic resources for improving these traits via marker-assisted selection (MAS).

Dissecting Key Adaptation Traits in the Polyploid Perennial Medicago sativa Using GBS-SNP Mapping.

Understanding key adaptation traits is crucial to developing new cultivars with broad adaptations. The main objective of this research is to understand the genetic basis of winter hardiness (WH) and fall dormancy (FD) in alfalfa and the association between the two traits. QTL analysis was conducted in a pseudo-testcross F1 population developed from two cultivars contrasting in FD (3010 with FD = 2 and CW 1010 with FD = 10). The mapping population was evaluated in three replications at two locations (Watkinsville and Blairsville, GA). FD levels showed low to moderate correlations with WH (0.22-0.57). Assessing dormancy in winter is more reliable than in the fall in southern regions with warm winters. The mapping population was genotyped using Genotyping-by-sequencing (GBS). Single dose allele SNPs (SDA) were used for constructing linkage maps. The parental map (CW 1010) consisted of 32 linkage groups spanning 2127.5 cM with 1377 markers and an average marker density of 1.5 cM/SNP. The maternal map (3010) had 32 linkage groups spanning 2788.4 cM with 1837 SDA SNPs with an average marker density of 1.5 cM/SNP. Forty-five significant (P < 0.05) QTLs for FD and 35 QTLs for WH were detected on both male and female linkage maps. More than 75% (22/28) of the dormancy QTL detected from the 3010 parent did not share genomic regions with WH QTLs and more than 70% (12/17) dormancy QTLs detected from CW 1010 parent were localized in different genomic regions than WH QTLs. These results suggest that the two traits have independent inheritance and therefore can be improved separately in breeding programs.

Insights into the Drought and Heat Avoidance Mechanism in Summer-Dormant Mediterranean Tall Fescue.

Summer dormancy is an evolutionary response that some perennial cool-season grasses adopted as an avoidance strategy to escape summer drought and heat. It is correlated with superior survival after severe summer droughts in many perennial grass species originating from Mediterranean environments. Understanding the genetic mechanism and environmental determinants of summer dormancy is important for interpreting the evolutionary history of seasonal dormancy and for the development of genomic tools to improve the efficiency of genetic selection for this important trait. The objectives of this research are to assess morphological and biochemical attributes that seem to be specific for the characterization of summer dormancy in tall fescue, and to validate the hypothesis that genes underlying stem determinacy might be involved in the mechanism of summer dormancy. Our results suggest that vernalization is an important requirement in the onset of summer dormancy in tall fescue. Non-vernalized tall fescue plants do not exhibit summer dormancy as vernalized plants do and behave more like summer-active types. This is manifested by continuation of shoot growth and high root activity in water uptake during summer months. Therefore, summer dormancy in tall fescue should be tested only in plants that underwent vernalization and are not subjected to water deficit during summer months. Total phenolic concentration in tiller bases (antioxidants) does not seem to be related to vernalization. It is most likely an environmental response to protect meristems from oxidative stress. Sequence analysis of the TFL1 homolog CEN gene from tall fescue genotypes belonging to summer-dormant and summer-active tall fescue types showed a unique deletion of three nucleotides specific to the dormant genotypes. Higher tiller bud numbers in dormant plants that were not allowed to flower and complete the reproductive cycle, confirmed that stem determinacy is a major component in the mechanism of summer dormancy. The number of variables identified in these studies as potential players in summer dormancy in tall fescue including vernalization, TFL1/CEN, water status, and protection from oxidative stress are a further confirmation that summer dormancy is a quantitative trait controlled by several genes with varying effects and prone to genotype by environment interactions.