Genetic Basis Of Natural Variation Research Articles

Deciphering the Genetic Basis of Kernel Composition in a Maize Association Panel.

The primary objective in contemporary maize breeding is to pursue high quality alongside high yield. Deciphering the genetic basis of natural variation in starch, protein, oil, and fiber contents is essential for manipulating kernel composition, thereby enhancing the kernel quality and meeting growing demands. Here, we identified 12 to 88 statistically significant loci associated with kernel composition traits through a genome-wide association study (GWAS) using a panel of 212 diverse inbred lines. A regional association study pinpointed numerous causal candidate genes at these loci. Coexpression and protein-protein interaction network analyses of candidate genes revealed several causal genes directly or indirectly involved in the metabolic processes related to kernel composition traits. Subsequent mutant experiment revealed that nonsense mutations in ZmTIFY12 affect starch, protein, and fiber content, whereas nonsense mutations in ZmTT12 affect starch, protein, and oil content. These findings provide valuable guidance for improving kernel quality in maize breeding efforts.

Genetic basis of nectar guide trichome variation between bumblebee- and self-pollinated monkeyflowers (Mimulus): role of the MIXTA-like gene GUIDELESS

Nectar guide trichomes play crucial ecological roles in bee-pollinated flowers, as they serve as footholds and guides for foraging bees to access the floral rewards. However, the genetic basis of natural variation in nectar guide trichomes among species remains poorly understood. In this study, we performed genetic analysis of nectar guide trichome variation between two closely related monkeyflower (Mimulus) species, the bumblebee-pollinated Mimulus lewisii and self-pollinated M. parishii. We demonstrate that a MIXTA-like R2R3-MYB gene, GUIDELESS, is a major contributor to the nectar guide trichome length variation between the two species. The short-haired M. parishii carries a recessive allele due to non-synonymous substitutions in a highly conserved motif among MIXTA-like MYB proteins. Furthermore, our results suggest that besides GUIDELESS, additional loci encoding repressors of trichome elongation also contribute to the transition from bumblebee-pollination to selfing. Taken together, these results suggest that during a pollination syndrome switch, changes in seemingly complex traits such as nectar guide trichomes could have a relatively simple genetic basis, involving just a few genes of large effects.

Natural variation of an autophagy-family gene among rice subspecies affects grain size and weight

Elucidating the genetic basis of natural variation in grain size and weight among rice varieties can help breeders develop high-yielding varieties. We identified a novel gene, GW3a (Grain Weight 3a) (LOC_Os03g27350), that affects rice grain size and weight. gw3a mutants showed higher total starch content and dry matter accumulation than the wild type (WT), Nipponbare, suggesting that GW3a negatively regulates grain size and weight. Moreover, our study found that GW3a interacted with OsATG8 by cleaving it, suggesting that GW3a may be involved in the assembly of autophagosomes and starch degradation in plants. The haplotype analysis of GW3a showed functional differences between indica and japonica rice. Taken together, we conclude that GW3a is expressed in the autophagosome pathway regulating starch metabolism in rice, affecting yield-related traits, such as grain size, grain weight and thousand grain weight (TGW). Our findings also shed new light on autophagy-mediated yield trait regulation, proposing a possible strategy for the genetic improvement of high-yield germplasm in rice.

Natural variation in CYCLIC NUCLEOTIDE-GATED ION CHANNEL 4 reveals a novel role of calcium signaling in vegetative phase change in Arabidopsis.

The timing of vegetative phase change (VPC) in plants is regulated by a temporal decline in the expression of miR156. Both exogenous cues and endogenous factors, such as temperature, light, sugar, nutrients, and epigenetic regulators, have been shown to affect VPC by altering miR156 expression. However, the genetic basis of natural variation in VPC remains largely unexplored. Here, we conducted a genome-wide association study on the variation of the timing of VPC in Arabidopsis. We identified CYCLIC NUCLEOTIDE-GATED ION CHANNEL 4 (CNGC4) as a significant locus associated with the diversity of VPC. Mutations in CNGC4 delayed VPC, accompanied by an increased expression level of miR156 and a corresponding decrease in SQUAMOSA PROMOTER BINDING-LIKE (SPL) gene expression. Furthermore, mutations in CNGC2 and CATION EXCHANGER 1/3 (CAX1/3) also led to a delay in VPC. Polymorphisms in the CNGC4 promoter contribute to the natural variation in CNGC4 expression and the diversity of VPC. Specifically, the early CNGC4 variant promotes VPC and enhances plant adaptation to local environments. In summary, our findings offer genetic insights into the natural variation in VPC in Arabidopsis, and reveal a previously unidentified role of calcium signaling in the regulation of VPC.

Natural polymorphisms in ZmIRX15A affect water-use efficiency by modulating stomatal density in maize.

Stomatal density (SD) is closely related to crop drought resistance. Understanding the genetic basis for natural variation in SD may facilitate efforts to improve water-use efficiency. Here, we report a genome-wide association study for SD in maize seedlings, which identified 18 genetic variants that could be resolved to seven candidate genes. A 3-bp insertion variant (InDel1089) in the last exon of Zea mays (Zm) IRX15A (Irregular xylem 15A) had the most significant association with SD and modulated the translation of ZmIRX15A mRNA by affecting its secondary structure. Dysfunction of ZmIRX15A increased SD, leading to an increase in the transpiration rate and CO2 assimilation efficiency. ZmIRX15A encodes a xylan deposition enzyme and its disruption significantly decreased xylan abundance in secondary cell wall composition. Transcriptome analysis revealed a substantial alteration of the expression of genes involved in stomatal complex morphogenesis and drought response in the loss-of-function of ZmIRX15A mutant. Overall, our study provides important genetic insights into the natural variation of leaf SD in maize, and the identified loci or genes can serve as direct targets for enhancing drought resistance in molecular-assisted maize breeding.

GWAS of grain color and tannin content in Chinese sorghum based on whole-genome sequencing

Key messageSeventy-three QTL related to grain color and tannin content were identified in Chinese sorghum accessions, and a new recessive allelic variant of TAN2 gene was discovered.Sorghum is mainly used for brewing distilled liquors in China. Since grain tannins play an important role in liquor brewing, accurately understanding the relationship between grain color and tannin content can provide basis for selection standards of tannin sorghum. We resequenced a panel of 242 Chinese sorghum accessions and performed population structure and genome-wide association study (GWAS) to identify quantitative trait locus (QTL) affecting pericarp color, testa pigment, and tannin content. Phylogenetic analysis, principal component analysis (PCA), and admixture model were used to infer population structure. Two distinct genetic sub-populations were identified according to their corresponding northern and southern geographic origin. To investigate the genetic basis of natural variation in sorghum grain color, GWAS with 2,760,264 SNPs was conducted in four environments using multiple models (Blink, FarmCPU, GLM, and MLM). Seventy-three QTL were identified to be associated for the color of exocarp, mesocarp, testa, and tannin content on all chromosomes except chromosome 5, of which 47 might be novel QTL. Some important QTL were found to colocalize with orthologous genes in the flavonoid biosynthetic pathway from other plants, including orthologous of Arabidopsis (Arabidopsis thaliana) TT2, TT7, TT12, TT16 and AT5G41220 (GST), as well as orthologous of rice (Oryza sativa) MYB61 and OsbHLH025. Our investigation of the variation in grain color and tannin content in Chinese sorghum germplasm may help guide future sorghum breeding for liquor brewing.

Natural variation and domestication selection of ZmSULTR3;4 is associated with maize lateral root length in response to salt stress.

Soil salinity is a major constraint that restricts crop productivity worldwide. Lateral roots (LRs) are important for water and nutrient acquisition, therefore understanding the genetic basis of natural variation in lateral root length (LRL) is of great agronomic relevance to improve salt tolerance in cultivated germplasms. Here, using a genome-wide association study, we showed that the genetic variation in ZmSULTR3;4, which encodes a plasma membrane-localized sulfate transporter, is associated with natural variation in maize LRL under salt stress. The transcript of ZmSULTR3;4 was found preferentially in the epidermal and vascular tissues of root and increased by salt stress, supporting its essential role in the LR formation under salt stress. Further candidate gene association analysis showed that DNA polymorphisms in the promoter region differentiate the expression of ZmSULTR3;4 among maize inbred lines that may contribute to the natural variation of LRL under salt stress. Nucleotide diversity and neutrality tests revealed that ZmSULTR3;4 has undergone selection during maize domestication and improvement. Overall, our results revealed a regulatory role of ZmSULTR3;4 in salt regulated LR growth and uncovered favorable alleles of ZmSULTR3;4, providing an important selection target for breeding salt-tolerant maize cultivar.

The genetic basis of natural variation in the timing of vegetative phase change in Arabidopsis thaliana.

The juvenile-to-adult transition in plants is known as vegetative phase change and is marked by changes in the expression of leaf traits in response to a decrease in the level of miR156 and miR157. To determine whether this is the only mechanism of vegetative phase change, we measured the appearance of phase-specific leaf traits in 70 natural accessions of Arabidopsis thaliana. We found that leaf shape was poorly correlated with abaxial trichome production (two adult traits), that variation in these traits was not necessarily correlated with the level of miR156, and that there was little to no correlation between the appearance of adult-specific vegetative traits and flowering time. We identified eight quantitative trait loci controlling phase-specific vegetative traits from a cross between the Columbia (Col-0) and Shakdara (Sha) accessions. Only one of these quantitative trait loci includes genes known to regulate vegetative phase change (MIR156A and TOE1), which were expressed at levels consistent with the precocious phenotype of Sha. Our results suggest that vegetative phase change is regulated both by the miR156/SPL module and by genes specific to different vegetative traits, and that natural variation in vegetative phase change can arise from either source.

QTL mapping and candidate gene analysis of low temperature germination in rice (Oryza sativa L.) using a genome wide association study.

Low temperature germination (LTG) is a key agronomic trait in rice (Oryza sativa L.). However, the genetic basis of natural variation for LTG is largely unknown. Here, a genome-wide association study (GWAS) was performed using 276 accessions from the 3,000 Rice Genomes (3K-RG) project with 497 k single nucleotide polymorphisms (SNPs) to uncover potential genes for LTG in rice. In total, 37 quantitative trait loci (QTLs) from the 6th day (D6) to the 10th day (D10) were detected in the full population, overlapping with 12 previously reported QTLs for LTG. One novel QTL, namely qLTG1-2, was found stably on D7 in both 2019 and 2020. Based on two germination-specific transcriptome datasets, 13 seed-expressed genes were isolated within a 200 kb interval of qLTG1-2. Combining with haplotype analysis, a functional uncharacterized gene, LOC_Os01g23580, and a seed germination-associated gene, LOC_Os01g23620 (OsSar1a), as promising candidate genes, both of which were significantly differentially expressed between high and low LTG accessions. Collectively, the candidate genes with favorable alleles may be useful for the future characterization of the LTG mechanism and the improvement of the LTG trait in rice breeding.

Genetic Incorporation of the Favorable Alleles for Three Genes Associated With Spikelet Development in Wheat.

The number of spikelets per spike is an important trait that directly affects grain yield in wheat. Three quantitative trait loci (QTLs) associated with spikelet nodes per spike (SNS) were mapped in a population of recombinant inbred lines generated from a cross between two advanced breeding lines of winter wheat based on the phenotypic variation evaluated over six locations/years. Two of the three QTLs are QSns.sxau-2A at the WHEATFRIZZY PANICLE (WFZP) loci and QSns.sxau-7A at the WHEAT ORTHOLOG OF APO1 (WAPO1) loci. The WFZP-A1b allele with a 14-bp deletion at QSns.sxau-2A was associated with increased spikelets per spike. WAPO-A1e, as a novel allele at WAPO1, were regulated at the transcript level that was associated with the SNS trait. The third SNS QTL, QSns.sxau-7D on chromosome 7D, was not associated with homoeologous WAPO-D1 or any other genes known to regulate SNS. The favorable alleles for each of WZFP-A1, WAPO-A1, and QSns.sxau-7D are identified and incorporated to increase up to 3.4 spikelets per spike in the RIL lines. Molecular markers for the alleles were developed. This study has advanced our understanding of the genetic basis of natural variation in spikelet development in wheat.

Multi-omics analysis dissects the genetic architecture of seed coat content in Brassica napus

BackgroundBrassica napus is an important vegetable oil source worldwide. Seed coat content is a complex quantitative trait that negatively correlates with the seed oil content in B. napus.ResultsHere we provide insights into the genetic basis of natural variation of seed coat content by transcriptome-wide association studies (TWAS) and genome-wide association studies (GWAS) using 382 B. napus accessions. By population transcriptomic analysis, we identify more than 700 genes and four gene modules that are significantly associated with seed coat content. We also characterize three reliable quantitative trait loci (QTLs) controlling seed coat content by GWAS. Combining TWAS and correlation networks of seed coat content-related gene modules, we find that BnaC07.CCR-LIKE (CCRL) and BnaTT8s play key roles in the determination of the trait by modulating lignin biosynthesis. By expression GWAS analysis, we identify a regulatory hotspot on chromosome A09, which is involved in controlling seed coat content through BnaC07.CCRL and BnaTT8s. We then predict the downstream genes regulated by BnaTT8s using multi-omics datasets. We further experimentally validate that BnaCCRL and BnaTT8 positively regulate seed coat content and lignin content. BnaCCRL represents a novel identified gene involved in seed coat development. Furthermore, we also predict the key genes regulating carbon allocation between phenylpropane compounds and oil during seed development in B. napus.ConclusionsThis study helps us to better understand the complex machinery of seed coat development and provides a genetic resource for genetic improvement of seed coat content in B. napus breeding.

The Barley and Wheat Pan-Genomes.

To unlock the genetic potential in crops, multi-genome comparisons are an essential tool. Decreasing costs and improved sequencing technologies have democratized plant genome sequencing and led to a vast increase in the amount of available reference sequences on the one hand and enabled the assembly of even the largest and most complex and repetitive crops genomes such as wheat and barley. These developments have led to the era of pan-genomics in recent years. Pan-genome projects enable the definition of the core and dispensable genome for various crop species as well as the analysis of structural and functional variation and hence offer unprecedented opportunities for exploring and utilizing the genetic basis of natural variation in crops. Comparing, analyzing, and visualizing these multiple reference genomes and their diversity requires powerful and specialized computational strategies and tools.

Genome-wide association study reveals the genetic architecture of root hair length in maize

BackgroundRoot hair, a special type of tubular-shaped cell, outgrows from root epidermal cell and plays important roles in the acquisition of nutrients and water, as well as interactions with biotic and abiotic stress. Although many genes involved in root hair development have been identified, genetic basis of natural variation in root hair growth has never been explored.ResultsHere, we utilized a maize association panel including 281 inbred lines with tropical, subtropical, and temperate origins to decipher the phenotypic diversity and genetic basis of root hair length. We demonstrated significant associations of root hair length with many metabolic pathways and other agronomic traits. Combining root hair phenotypes with 1.25 million single nucleotide polymorphisms (SNPs) via genome-wide association study (GWAS) revealed several candidate genes implicated in cellular signaling, polar growth, disease resistance and various metabolic pathways.ConclusionsThese results illustrate the genetic basis of root hair length in maize, offering a list of candidate genes predictably contributing to root hair growth, which are invaluable resource for the future functional investigation.

Phenotyping stomatal closure by thermal imaging for GWAS and TWAS of water use efficiency-related genes.

Stomata allow CO2 uptake by leaves for photosynthetic assimilation at the cost of water vapor loss to the atmosphere. The opening and closing of stomata in response to fluctuations in light intensity regulate CO2 and water fluxes and are essential for maintaining water-use efficiency (WUE). However, a little is known about the genetic basis for natural variation in stomatal movement, especially in C4 crops. This is partly because the stomatal response to a change in light intensity is difficult to measure at the scale required for association studies. Here, we used high-throughput thermal imaging to bypass the phenotyping bottleneck and assess 10 traits describing stomatal conductance (gs) before, during and after a stepwise decrease in light intensity for a diversity panel of 659 sorghum (Sorghum bicolor) accessions. Results from thermal imaging significantly correlated with photosynthetic gas exchange measurements. gs traits varied substantially across the population and were moderately heritable (h2 up to 0.72). An integrated genome-wide and transcriptome-wide association study identified candidate genes putatively driving variation in stomatal conductance traits. Of the 239 unique candidate genes identified with the greatest confidence, 77 were putative orthologs of Arabidopsis (Arabidopsis thaliana) genes related to functions implicated in WUE, including stomatal opening/closing (24 genes), stomatal/epidermal cell development (35 genes), leaf/vasculature development (12 genes), or chlorophyll metabolism/photosynthesis (8 genes). These findings demonstrate an approach to finding genotype-to-phenotype relationships for a challenging trait as well as candidate genes for further investigation of the genetic basis of WUE in a model C4 grass for bioenergy, food, and forage production.

Genetic basis of kernel starch content decoded in a maize multi-parent population.

SummaryStarch is the most abundant storage carbohydrate in maize kernels and provides calories for humans and other animals as well as raw materials for various industrial applications. Decoding the genetic basis of natural variation in kernel starch content is needed to manipulate starch quantity and quality via molecular breeding to meet future needs. Here, we identified 50 unique single quantitative trait loci (QTLs) for starch content with 18 novel QTLs via single linkage mapping, joint linkage mapping and a genome‐wide association study in a multi‐parent population containing six recombinant inbred line populations. Only five QTLs explained over 10% of phenotypic variation in single populations. In addition to a few large‐effect and many small‐effect additive QTLs, limited pairs of epistatic QTLs also contributed to the genetic basis of the variation in kernel starch content. A regional association study identified five non‐starch‐pathway genes that were the causal candidate genes underlying the identified QTLs for starch content. The pathway‐driven analysis identified ZmTPS9, which encodes a trehalose‐6‐phosphate synthase in the trehalose pathway, as the causal gene for the QTL qSTA4−2, which was detected by all three statistical analyses. Knockout of ZmTPS9 increased kernel starch content and, in turn, kernel weight in maize, suggesting potential applications for ZmTPS9 in maize starch and yield improvement. These findings extend our knowledge about the genetic basis of starch content in maize kernels and provide valuable information for maize genetic improvement of starch quantity and quality.

High-resolution genome-wide association study pinpoints metal transporter and chelator genes involved in the genetic control of element levels in maize grain.

Despite its importance to plant function and human health, the genetics underpinning element levels in maize grain remain largely unknown. Through a genome-wide association study in the maize Ames panel of nearly 2,000 inbred lines that was imputed with ∼7.7 million SNP markers, we investigated the genetic basis of natural variation for the concentration of 11 elements in grain. Novel associations were detected for the metal transporter genes rte2 (rotten ear2) and irt1 (iron-regulated transporter1) with boron and nickel, respectively. We also further resolved loci that were previously found to be associated with one or more of five elements (copper, iron, manganese, molybdenum, and/or zinc), with two metal chelator and five metal transporter candidate causal genes identified. The nas5 (nicotianamine synthase5) gene involved in the synthesis of nicotianamine, a metal chelator, was found associated with both zinc and iron and suggests a common genetic basis controlling the accumulation of these two metals in the grain. Furthermore, moderate predictive abilities were obtained for the 11 elemental grain phenotypes with two whole-genome prediction models: Bayesian Ridge Regression (0.33–0.51) and BayesB (0.33–0.53). Of the two models, BayesB, with its greater emphasis on large-effect loci, showed ∼4–10% higher predictive abilities for nickel, molybdenum, and copper. Altogether, our findings contribute to an improved genotype-phenotype map for grain element accumulation in maize.

Natural Variation in Crops: Realized Understanding, Continuing Promise.

Crops feed the world's population and shape human civilization. The improvement of crop productivity has been ongoing for almost 10,000 years and has evolved from an experience-based to a knowledge-driven practice over the past three decades. Natural alleles and their reshuffling are long-standing genetic changes that affect how crops respond to various environmental conditions and agricultural practices. Decoding the genetic basis of natural variation is central to understanding crop evolution and, in turn, improving crop breeding. Here, we review current advances in the approaches used to map the causal alleles of natural variation, provide refined insights into the genetics and evolution of natural variation, and outline how this knowledge promises to drive the development of sustainable agriculture under the dome of emerging technologies.

Genome- and transcriptome-wide association studies provide insights into the genetic basis of natural variation of seed oil content in Brassica napus

Seed oil content (SOC) is a highly important and complex trait in oil crops. Here, we decipher the genetic basis of natural variation in SOC of Brassica napus by genome- and transcriptome-wide association studies using 505 inbred lines. We mapped reliable quantitative trait loci (QTLs) that control SOC in eight environments, evaluated the effect of each QTL on SOC, and analyzed selection in QTL regions during breeding. Six-hundred and ninety-two genes and four gene modules significantly associated with SOC were identified by analyzing population transcriptomes from seeds. A gene prioritization framework, POCKET (prioritizing the candidate genes by incorporating information on knowledge-based gene sets, effects of variants, genome-wide association studies, and transcriptome-wide association studies), was implemented to determine the causal genes in the QTL regions based on multi-omic datasets. A pair of homologous genes, BnPMT6s, in two QTLs were identified and experimentally demonstrated to negatively regulate SOC. This study provides rich genetic resources for improving SOC and valuable insights toward understanding the complex machinery that directs oil accumulation in the seeds of B. napus and other oil crops.

Genetic Basis of Natural Variation in Spontaneous Grooming in Drosophila melanogaster.

Spontaneous grooming behavior is a component of insect fitness. We quantified spontaneous grooming behavior in 201 sequenced lines of the Drosophila melanogaster Genetic Reference Panel and observed significant genetic variation in spontaneous grooming, with broad-sense heritabilities of 0.25 and 0.24 in females and males, respectively. Although grooming behavior is highly correlated between males and females, we observed significant sex by genotype interactions, indicating that the genetic basis of spontaneous grooming is partially distinct in the two sexes. We performed genome-wide association analyses of grooming behavior, and mapped 107 molecular polymorphisms associated with spontaneous grooming behavior, of which 73 were in or near 70 genes and 34 were over 1 kilobase from the nearest gene. The candidate genes were associated with a wide variety of gene ontology terms, and several of the candidate genes were significantly enriched in a genetic interaction network. We performed functional assessments of 29 candidate genes using RNA interference, and found that 11 affected spontaneous grooming behavior. The genes associated with natural variation in Drosophila grooming are involved with glutamate metabolism (Gdh) and transport (Eaat); interact genetically with (CCKLR-17D1) or are in the same gene family as (PGRP-LA) genes previously implicated in grooming behavior; are involved in the development of the nervous system and other tissues; or regulate the Notch and Epidermal growth factor receptor signaling pathways. Several DGRP lines exhibited extreme grooming behavior. Excessive grooming behavior can serve as a model for repetitive behaviors diagnostic of several human neuropsychiatric diseases.

Genetic architecture underlying changes in carotenoid accumulation during the evolution of the blind Mexican cavefish, Astyanax mexicanus.

Carotenoids are lipid-soluble yellow to orange pigments produced by plants, bacteria, and fungi. They are consumed by animals and metabolized to produce molecules essential for gene regulation, vision, and pigmentation. Cave animals represent an interesting opportunity to understand how carotenoid utilization evolves. Caves are devoid of light, eliminating primary production of energy through photosynthesis and, therefore, limiting carotenoid availability. Moreover, the selective pressures that favor carotenoid-based traits, like pigmentation and vision, are relaxed. Astyanax mexicanus is a species of fish with multiple river-adapted (surface) and cave-adapted populations (i.e., Tinaja, Pachón, Molino). Cavefish exhibit regressive features, such as loss of eyes and melanin pigment, and constructive traits, like increased sensory neuromasts and starvation resistance. Here, we show that, unlike surface fish, Tinaja and Pachón cavefish accumulate carotenoids in the visceral adipose tissue. Carotenoid accumulation is not observed in Molino cavefish, indicating that it is not an obligatory consequence of eye loss. We used quantitative trait loci mapping and RNA sequencing to investigate genetic changes associated with carotenoid accumulation. Our findings suggest that multiple stages of carotenoid processing may be altered in cavefish, including absorption and transport of lipids, cleavage of carotenoids into unpigmented molecules, and differential development of intestinal cell types involved in carotenoid assimilation. Our study establishes A. mexicanus as a model to study the genetic basis of natural variation in carotenoid accumulation and how it impacts physiology.