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Identification of Candidate Genes Regulating Drought Tolerance in Pearl Millet.

ABSTRACT: Pearl millet is an important crop of the arid and semi-arid ecologies to sustain food and fodder production. The greater tolerance to drought stress attracts us to examine its cellular and molecular mechanisms via functional genomics approaches to augment the grain yield. Here, we studied the drought response of 48 inbreds representing four different maturity groups at the flowering stage. A set of 74 drought-responsive genes were separated into five major phylogenic groups belonging to eight functional groups, namely ABA signaling, hormone signaling, ion and osmotic homeostasis, TF-mediated regulation, molecular adaptation, signal transduction, physiological adaptation, detoxification, which were comprehensively studied. Among the conserved motifs of the drought-responsive genes, the protein kinases and MYB domain proteins were the most conserved ones. Comparative in-silico analysis of the drought genes across millet crops showed foxtail millet had most orthologs with pearl millet. Of 698 haplotypes identified across millet crops, MyC2 and Myb4 had maximum haplotypes. The protein-protein interaction network identified ABI2, P5CS, CDPK, DREB, MYB, and CYP707A3 as major hub genes. The expression assay showed the presence of common as well as unique drought-responsive genes across maturity groups. Drought tolerant genotypes in respective maturity groups were identified from the expression pattern of genes. Among several gene families, ABA signaling, TFs, and signaling proteins were the prospective contributors to drought tolerance across maturity groups. The functionally validated genes could be used as promising candidates in backcross breeding, genomic selection, and gene-editing schemes in pearl millet and other millet crops to increase the yield in drought-prone arid and semi-arid ecologies.

SUBMITTER: Chakraborty A

PROVIDER: S-EPMC9266394 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundIdentification of genes underlying drought tolerance (DT) quantitative trait loci (QTLs) will facilitate understanding of molecular mechanisms of drought tolerance, and also will accelerate genetic improvement of pearl millet through marker-assisted selection. We report a map based on genes with assigned functional roles in plant adaptation to drought and other abiotic stresses and demonstrate its use in identifying candidate genes underlying a major DT-QTL.ResultsSeventy five single nucleotide polymorphism (SNP) and conserved intron spanning primer (CISP) markers were developed from available expressed sequence tags (ESTs) using four genotypes, H 77/833-2, PRLT 2/89-33, ICMR 01029 and ICMR 01004, representing parents of two mapping populations. A total of 228 SNPs were obtained from 30.5 kb sequenced region resulting in a SNP frequency of 1/134 bp. The positions of major pearl millet linkage group (LG) 2 DT-QTLs (reported from crosses H 77/833-2 × PRLT 2/89-33 and 841B × 863B) were added to the present consensus function map which identified 18 genes, coding for PSI reaction center subunit III, PHYC, actin, alanine glyoxylate aminotransferase, uridylate kinase, acyl-CoA oxidase, dipeptidyl peptidase IV, MADS-box, serine/threonine protein kinase, ubiquitin conjugating enzyme, zinc finger C- × 8-C × 5-C × 3-H type, Hd3, acetyl CoA carboxylase, chlorophyll a/b binding protein, photolyase, protein phosphatase1 regulatory subunit SDS22 and two hypothetical proteins, co-mapping in this DT-QTL interval. Many of these candidate genes were found to have significant association with QTLs of grain yield, flowering time and leaf rolling under drought stress conditions.ConclusionsWe have exploited available pearl millet EST sequences to generate a mapped resource of seventy five new gene-based markers for pearl millet and demonstrated its use in identifying candidate genes underlying a major DT-QTL in this species. The reported gene-based markers represent an important resource for identification of candidate genes for other mapped abiotic stress QTLs in pearl millet. They also provide a resource for initiating association studies using candidate genes and also for comparing the structure and function of distantly related plant genomes such as other Poaceae members.

Project description:Pearl millet is a nutri-cereal that is mostly grown in harsh environments, making it an ideal crop to study heat tolerance mechanisms at the molecular level. Despite having a better-inbuilt tolerance to high temperatures than other crops, heat stress negatively affects the crop, posing a threat to productivity gain. Hence, to understand the heat-responsive genes, the leaf and root samples of two contrasting pearl millet inbreds, EGTB 1034 (heat tolerant) and EGTB 1091 (heat sensitive), were subjected to heat-treated conditions and generated genome-wide transcriptomes. We discovered 13,464 differentially expressed genes (DEGs), of which 6932 were down-regulated and 6532 up-regulated in leaf and root tissues. The pairwise analysis of the tissue-based transcriptome data of the two genotypes demonstrated distinctive genotype and tissue-specific expression of genes. The root exhibited a higher number of DEGs compared to the leaf, emphasizing different adaptive strategies of pearl millet. A large number of genes encoding ROS scavenging enzymes, WRKY, NAC, enzymes involved in nutrient uptake, protein kinases, photosynthetic enzymes, and heat shock proteins (HSPs) and several transcription factors (TFs) involved in cross-talking of temperature stress responsive mechanisms were activated in the stress conditions. Ribosomal proteins emerged as pivotal hub genes, highly interactive with key genes expressed and involved in heat stress response. The synthesis of secondary metabolites and metabolic pathways of pearl millet were significantly enriched under heat stress. Comparative synteny analysis of HSPs and TFs in the foxtail millet genome demonstrated greater collinearity with pearl millet compared to proso millet, rice, sorghum, and maize. In this study, 1906 unannotated DEGs were identified, providing insight into novel participants in the molecular response to heat stress. The identified genes hold promise for expediting varietal development for heat tolerance in pearl millet and similar crops, fostering resilience and enhancing grain yield in heat-prone environments.

Project description:A pearl millet inbred germplasm association panel (PMiGAP) comprising 250 inbred lines, representative of cultivated germplasm from Africa and Asia, elite improved open-pollinated cultivars, hybrid parental inbreds and inbred mapping population parents, was recently established. This study presents the first report of genetic diversity in PMiGAP and its exploitation for association mapping of drought tolerance traits. For diversity and genetic structure analysis, PMiGAP was genotyped with 37 SSR and CISP markers representing all seven linkage groups. For association analysis, it was phenotyped for yield and yield components and morpho-physiological traits under both well-watered and drought conditions, and genotyped with SNPs and InDels from seventeen genes underlying a major validated drought tolerance (DT) QTL. The average gene diversity in PMiGAP was 0.54. The STRUCTURE analysis revealed six subpopulations within PMiGAP. Significant associations were obtained for 22 SNPs and 3 InDels from 13 genes under different treatments. Seven SNPs associations from 5 genes were common under irrigated and one of the drought stress treatments. Most significantly, an important SNP in putative acetyl CoA carboxylase gene showed constitutive association with grain yield, grain harvest index and panicle yield under all treatments. An InDel in putative chlorophyll a/b binding protein gene was significantly associated with both stay-green and grain yield traits under drought stress. This can be used as a functional marker for selecting high yielding genotypes with 'stay green' phenotype under drought stress. The present study identified useful marker-trait associations of important agronomics traits under irrigated and drought stress conditions with genes underlying a major validated DT-QTL in pearl millet. Results suggest that PMiGAP is a useful panel for association mapping. Expression patterns of genes also shed light on some physiological mechanisms underlying pearl millet drought tolerance.

Project description:Water deficit stress at the early stage of development is one of the main factors limiting pearl millet production. One practice to counteract this limitation would be to resort to the application of hormones to stimulate plant growth and development at critical stages. Exogenous methyl jasmonate (MeJA) can improve drought tolerance by modulating signaling, metabolism, and photosynthesis pathways, therefore, we assumed that can occur in pearl millet during the early stage of development. To decipher the molecular mechanisms controlling these pathways, RNAseq was conducted in two pearl millet genotypes, drought-sensitive SosatC88 and drought-tolerant Souna3, in response to 200 μM of MeJA. Pairwise comparison between the MeJA-treated and non-treated plants revealed 3270 differentially expressed genes (DEGs) among 20,783 transcripts in SosatC88 and 127 DEGs out of 20,496 transcripts in Souna3. Gene ontology (GO) classification assigned most regulated DEGs in SosatC88 to heme binding, oxidation-reduction process, response to oxidative stress and membrane, and in Souna3 to terpene synthase activity, lyase activity, magnesium ion binding, and thylakoid. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis reveals that DEGs in SosatC88 are related to the oxidation-reduction process, the biosynthesis of other secondary metabolites, the signal transduction, and the metabolism of terpenoids, while in Souna3, DEGs are related to the metabolism of terpenoids and the energy metabolism. Two genes encoding a diterpenoid biosynthesis-related (Pgl_GLEAN_10009413) and a Glutathione S transferase T3 (Pgl_GLEAN_10034098) were contra-regulated between SosatC88 and Souna3. Additionally, five random genes differentially expressed by RNAseq were validated using qPCR, therefore, they are potential targets for the development of novel strategies breeding schemes for plant growth under water deficit stress. These insights into the molecular mechanisms of pearl millet genotype tolerance at the early stage of development contribute to the understanding of the role of hormones in adaptation to drought-prone environments.

Project description:BACKGROUND: Drought stress is one of the major limiting factors for maize production. With the availability of maize B73 reference genome and whole-genome resequencing of 15 maize inbreds, common variants (CV) and clustering analyses were applied to identify non-synonymous SNPs (nsSNPs) and corresponding candidate genes for drought tolerance. RESULTS: A total of 524 nsSNPs that were associated with 271 candidate genes involved in plant hormone regulation, carbohydrate and sugar metabolism, signaling molecules regulation, redox reaction and acclimation of photosynthesis to environment were detected by CV and cluster analyses. Most of the nsSNPs identified were clustered in bin 1.07 region that harbored six previously reported QTL with relatively high phenotypic variation explained for drought tolerance. Genes Ontology (GO) analysis of candidate genes revealed that there were 35 GO terms related to biotic stimulus and membrane-bounded organelle, showing significant differences between the candidate genes and the reference B73 background. Changes of expression level in these candidate genes for drought tolerance were detected using RNA sequencing for fertilized ovary, basal leaf meristem tissue and roots collected under drought stressed and well-watered conditions. The results indicated that 70% of candidate genes showed significantly expression changes under two water treatments and our strategies for mining candidate genes are feasible and relatively efficient. CONCLUSIONS: Our results successfully revealed candidate nsSNPs and associated genes for drought tolerance by comparative sequence analysis of 16 maize inbred lines. Both methods we applied were proved to be efficient for identifying candidate genes for complex traits through the next-generation sequencing technologies (NGS). These selected genes will not only facilitate understanding of genetic basis of drought stress response, but also accelerate genetic improvement through marker-assisted selection in maize.

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Identification of Candidate Genes Regulating Drought Tolerance in Pearl Millet.

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