Integration of gene-based markers in a pearl millet genetic map for identification of candidate genes underlying drought tolerance QTLs

BACKGROUND: Identification 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. RESULTS: Seventy 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. CONCLUSIONS: We 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

Mapping Quantitative Trait Loci Controlling High Iron and Zinc Content in Self and Open Pollinated Grains of Pearl Millet [Pennisetum glaucum (L.) R. Br.]

Pearl millet is a multipurpose grain/fodder crop of the semi-arid tropics, feeding many of the world’s poorest and most undernourished people. Genetic variation among adapted pearl millet inbreds and hybrids suggests it will be possible to improve grain micronutrient concentrations by selective breeding. Using 305 loci, a linkage map was constructed to map QTLs for grain [Fe] and [Zn] using replicated samples of 106 pearl millet RILs (F6) derived from ICMB 841-P3 × 863B-P2. The grains of the RIL population were evaluated for Fe and Zn concentration using atomic absorption spectrophotometer. Grain mineral concentrations ranged from 28.4-124.0 ppm for Fe and 28.7-119.8 ppm for Zn. Similarly,grain Fe and Zn in open pollinated seeds ranged between 22.4-77.4 and 21.9-73.7 ppm, respectively. Mapping with 305 (96 SSRs; 208 DArT) markers detected seven linkage groups covering 1749 cM (Haldane) with an average intermarker distance of 5.73 cM. On the basis of two environment phenotypic data, two co-localized QTLs for Fe and Zn density on LG 3 were identified by composite interval mapping (CIM). Fe QTL explained 19% phenotypic variation, whereas the Zn QTL explained 36% phenotypic variation. Likewise for open pollinated seeds, the QTL analysis led to identification of two QTLs for grain Fe density on LG 3 and 5, and two QTLs for grain Zn density on LG 3 and 7. The total phenotypic variance for Fe and Zn QTLs in open pollinated seeds was 16% and 42%, respectively. Analysis of QTL × QTL and QTL × QTL× environment interactions indicated no major epistasis

Conventional and Molecular Breeding Approaches for Biofortification of Pearl Millet

Pearl millet [Pennisetum glaucum (L.) R. Br.] is an essential diet of more than 90 million people in the semi-arid tropics of the world where droughts and low fertility of soils cause frequent failures of other crops. It is an important nutri-rich grain cereal in the drier regions of the world grown on 26 mha by millions of farmers (IFAD 1999; Yadav and Rai 2013). This makes pearl millet the sixth most important crop in the world and fourth most important food crop of the India, next to rice, wheat, and maize with annual cultivation over an area of ~8 mha. Pearl millet is also primary food crop in sub-Saharan Africa and is grown on 15 mha (Yadav and Rai 2013). The significant increase in productivity of pearl millet in India is attributed to development and adoption of hybrids of early to medium duration maturity. More than 120 diverse hybrids/varieties have been released till date for various production environments. The heterosis breeding and improved crop management technologies increased productivity substantially achieving higher increased production of 9.80 mt in 2016–2017 from 2.60 mt in 1950–1951 in spite of declined of area under the crop by 20–30% over last two decades (Yadav et al. 2012)

Water saving traits co-map with a major terminal drought tolerance quantitative trait locus in pearl millet [Pennisetum glaucum (L.) R. Br.]

Low transpiration rates in pearl millet under fully irrigated conditions decrease plant water use at vegetative stage and then increase the water availability during grain filling and finally the terminal drought tolerance. Hundred and thirteen recombinant inbred lines developed from a cross between H77/833-2 and PRLT2/89-33 (terminal drought-sensitive × tolerant genotype) were evaluated to map transpiration rate (Tr, a proxy for canopy conductance), organ weights, leaf area and thickness and to study their interactions. Transpiration rate was increased by two H77/833-2 and two PRLT2/89-33 alleles on linkage group (LG) 2, whose importance depended on the vapor pressure deficit. The two H77/833-2 and one PRLT2/89-33 alleles co-mapped to a previously identified major terminal drought tolerance quantitative trait locus (QTL), although in a much smaller genetic interval. The other Tr allele from H77/833-2 also enhanced biomass dry weight and co-located with a formerly identified stover and tillering QTL. Leaf characteristics were linked to two loci on LG7. Plant water use was increased and decreased by different loci combinations for Tr, tillering and leaf characteristics, whose respective importance depended on the environmental conditions. Therefore, different alleles influence plant water use and have close interactions with one another and with the environment, so that different ideotypes for plant water use exist or could be designed from specific allele combinations conferring particular physiological characteristics for specific adaptation to a range of terminal drought conditions

Pearl millet [Pennisetum glaucum (L.) R. Br.] consensus linkage map constructed using four RIL mapping populations and newly developed EST-SSRs

Background: Pearl millet [Pennisetum glaucum (L.) R. Br.] is a widely cultivated drought-and high-temperature tolerant C4 cereal grown under dryland, rainfed and irrigated conditions in drought-prone regions of the tropics and sub-tropics of Africa, South Asia and the Americas. It is considered an orphan crop with relatively few genomic and genetic resources. This study was undertaken to increase the EST-based microsatellite marker and genetic resources for this crop to facilitate marker-assisted breeding. Results: Newly developed EST-SSR markers (99), along with previously mapped EST-SSR (17), genomic SSR (53) and STS (2) markers, were used to construct linkage maps of four F-7 recombinant inbred populations (RIP) based on crosses ICMB 841-P3 x 863B-P2 (RIP A), H 77/833-2 x PRLT 2/89-33 (RIP B), 81B-P6 x ICMP 451-P8 (RIP C) and PT 732B-P2 x P1449-2-P1 (RIP D). Mapped loci numbers were greatest for RIP A (104), followed by RIP B (78), RIP C (64) and RIP D (59). Total map lengths (Haldane) were 615 cM, 690 cM, 428 cM and 276 cM, respectively. A total of 176 loci detected by 171 primer pairs were mapped among the four crosses. A consensus map of 174 loci (899 cM) detected by 169 primer pairs was constructed using MergeMap to integrate the individual linkage maps. Locus order in the consensus map was well conserved for nearly all linkage groups. Eighty-nine EST-SSR marker loci from this consensus map had significant BLAST hits (top hits with e-value <= 1E-10) on the genome sequences of rice, foxtail millet, sorghum, maize and Brachypodium with 35, 88, 58, 48 and 38 loci, respectively. Conclusion: The consensus map developed in the present study contains the largest set of mapped SSRs reported to date for pearl millet, and represents a major consolidation of existing pearl millet genetic mapping information. This study increased numbers of mapped pearl millet SSR markers by >50%, filling important gaps in previously published SSR-based linkage maps for this species and will greatly facilitate SSR-based QTL mapping and applied marker-assisted selection programs.Keywords: Synteny, Linkage map, EST-SSR markers, Consensus map, EST, Pearl millet, Drought stres