Publications of NIPGR Scientists
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Item Genome-wide association study of major agronomic traits in foxtail millet (Setaria italica L.) using ddRAD sequencing(Springer Nature, 2019) Jaiswal, Vandana; Gupta, Sarika; Gahlaut, Vijay; Muthamilarasan, Mehanathan; Bandyopadhyay, Tirthankar; Ramchiary, Nirala; Prasad, ManojFoxtail millet (Setaria italica), the second largest cultivated millet crop after pearl millet, is utilized for food and forage globally. Further, it is also considered as a model crop for studying agronomic, nutritional and biofuel traits. In the present study, a genome-wide association study (GWAS) was performed for ten important agronomic traits in 142 foxtail millet core eco-geographically diverse genotypes using 10 K SNPs developed through GBS-ddRAD approach. Number of SNPs on individual chromosome ranged from 844 (chromosome 5) to 2153 (chromosome 8) with an average SNP frequency of 25.9 per Mb. The pairwise linkage disequilibrium (LD) estimated using the squared-allele frequency correlations was found to decay rapidly with the genetic distance of 177 Kb. However, for individual chromosome, LD decay distance ranged from 76 Kb (chromosome 6) to 357 Kb (chromosome 4). GWAS identified 81 MTAs (marker-trait associations) for ten traits across the genome. High confidence MTAs for three important agronomic traits including FLW (flag leaf width), GY (grain yield) and TGW (thousand-grain weight) were identified. Significant pyramiding effect of identified MTAs further supplemented its importance in breeding programs. Desirable alleles and superior genotypes identified in the present study may prove valuable for foxtail millet improvement through marker-assisted selection.Item Genome-wide association study (GWAS) delineates genomic loci for ten nutritional elements in foxtail millet (Setaria italica L.)(Elsevier B.V., 2019) Jaiswal, Vandana; Bandyopadhyay, Tirthankar; Gahlaut, Vijay; Gupta, Sarika; Dhaka, Annvi; Ramchiary, Nirala; Prasad, ManojNutritional deficiency is found to be a major threat to human health, especially in low-income countries. Thus it is essential to improve nutritional qualities of important food crops. Foxtail millet is second largest cultivated millet and 2–5 times nutritionally richer than major cereal crops. During the present study, we identified genetic determinants of ten nutritional elements including potassium, nickel, calcium, boron, magnesium, phosphorus, sulphur, zinc, manganese and iron for the first time in foxtail millet. For this purpose, genome-wide association studies (GWAS) were conducted using 93 diverse accessions and 10 K SNPs (distributed across all the nine foxtail millet chromosomes). Altogether, 74 marker-trait associations (MTAs) were identified to be associated with above mentioned ten elements, out of which ten (10) MTAs (associated with B, Mg, Zn and Fe) showed high confidence [-log(p) > 5.78]. Identified desirable SNP alleles and favourable haplotypes may prove useful in foxtail breeding. Also, significant pyramiding effect suggested that associated elements can be substantially enhanced through combining more than one MTA. Candidate genes residing within or near the association signal may be selected for functional characterization. Superior genotypes identified may prove as a potential donor in foxtail millet breeding assisted through the molecular marker.Item Population structure and association mapping of yield contributing agronomic traits in foxtail millet(Springer, 2014) Gupta, Sarika; Kumari, Kajal; Muthamilarasan, Mehanathan; Parida, Swarup K.; Prasad, ManojAssociation analyses accounting for population structure and relative kinship identified eight SSR markers ( p < 0.01) showing significant association ( R (2) = 18 %) with nine agronomic traits in foxtail millet. Association mapping is an efficient tool for identifying genes regulating complex traits. Although association mapping using genomic simple sequence repeat (SSR) markers has been successfully demonstrated in many agronomically important crops, very few reports are available on marker-trait association analysis in foxtail millet. In the present study, 184 foxtail millet accessions from diverse geographical locations were genotyped using 50 SSR markers representing the nine chromosomes of foxtail millet. The genetic diversity within these accessions was examined using a genetic distance-based and a general model-based clustering method. The model-based analysis using 50 SSR markers identified an underlying population structure comprising five sub-populations which corresponded well with distance-based groupings. The phenotyping of plants was carried out in the field for three consecutive years for 20 yield contributing agronomic traits. The linkage disequilibrium analysis considering population structure and relative kinship identified eight SSR markers (p < 0.01) on different chromosomes showing significant association (R (2) = 18 %) with nine agronomic traits. Four of these markers were associated with multiple traits. The integration of genetic and physical map information of eight SSR markers with their functional annotation revealed strong association of two markers encoding for phospholipid acyltransferase and ubiquitin carboxyl-terminal hydrolase located on the same chromosome (5) with flag leaf width and grain yield, respectively. Our findings on association mapping is the first report on Indian foxtail millet germplasm and this could be effectively applied in foxtail millet breeding to further uncover marker-trait associations with a large number of markers.Item Genome-wide development and use of microsatellite markers for large-scale genotyping applications in foxtail millet [Setaria italica (L.)](Oxford University Press, 2013) Pandey, Garima; Misra, Gopal; Kumari, Kajal; Gupta, Sarika; Parida, Swarup K.; Chattopadhyay, Debasis; Prasad, ManojThe availability of well-validated informative co-dominant microsatellite markers and saturated genetic linkage map has been limited in foxtail millet (Setaria italica L.). In view of this, we conducted a genome-wide analysis and identified 28 342 microsatellite repeat-motifs spanning 405.3 Mb of foxtail millet genome. The trinucleotide repeats (∼48%) was prevalent when compared with dinucleotide repeats (∼46%). Of the 28 342 microsatellites, 21 294 (∼75%) primer pairs were successfully designed, and a total of 15 573 markers were physically mapped on 9 chromosomes of foxtail millet. About 159 markers were validated successfully in 8 accessions of Setaria sp. with ∼67% polymorphic potential. The high percentage (89.3%) of cross-genera transferability across millet and non-millet species with higher transferability percentage in bioenergy grasses (∼79%, Switchgrass and ∼93%, Pearl millet) signifies their importance in studying the bioenergy grasses. In silico comparative mapping of 15 573 foxtail millet microsatellite markers against the mapping data of sorghum (16.9%), maize (14.5%) and rice (6.4%) indicated syntenic relationships among the chromosomes of foxtail millet and target species. The results, thus, demonstrate the immense applicability of developed microsatellite markers in germplasm characterization, phylogenetics, construction of genetic linkage map for gene/quantitative trait loci discovery, comparative mapping in foxtail millet, including other millets and bioenergy grass species.Item Foxtail millet: A model crop for genetic and genomic studies in bioenergy grasses(Informa UK Limited, an Informa Group Company (T&F Group), 2013) Lata, Charu; Gupta, Sarika; Prasad, ManojFoxtail millet is one of the oldest domesticated diploid C4 Panicoid crops having a comparatively small genome size of approximately 515 Mb, short life cycle, and inbreeding nature. Its two species, Setaria italica (domesticated) and Setaria viridis (wild progenitor), have characteristics that classify them as excellent model systems to examine several aspects of architectural, evolutionary, and physiological importance in Panicoid grasses especially the biofuel crops such as switchgrass and napiergrass. Foxtail millet is a staple crop used extensively for food and fodder in parts of Asia and Africa. In its long history of cultivation, it has been adapted to arid and semi-arid areas of Asia, North Africa, South and North America. Foxtail millet has one of the largest collections of cultivated as well as wild-type germplasm rich with phenotypic variations and hence provides prospects for association mapping and allele-mining of elite and novel variants to be incorporated in crop improvement programs. Most of the foxtail millet accessions can be primarily abiotic stress tolerant particularly to drought and salinity, and therefore exploiting these agronomic traits can enhance its efficacy in marker-aided breeding as well as in genetic engineering for abiotic stress tolerance. In addition, the release of draft genome sequence of foxtail millet would be useful to the researchers worldwide in not only discerning the molecular basis of biomass production in biofuel crops and the methods to improve it, but also for the introgression of beneficial agronomically important characteristics in foxtail millet as well as in related Panicoid bioenergy grasses.Item Development of eSSR-markers in Setaria italica and their applicability in studying genetic diversity, cross-transferability and comparative mapping in millet and non-millet species(PLOS, 2013) Kumari, Kajal; Muthamilarasan, Mehanathan; Misra, Gopal; Gupta, Sarika; Subramanian, Alagesan; Parida, Swarup K.; Chattopadhyay, Debasis; Prasad, ManojFoxtail millet (Setariaitalica L.) is a tractable experimental model crop for studying functional genomics of millets and bioenergy grasses. But the limited availability of genomic resources, particularly expressed sequence-based genic markers is significantly impeding its genetic improvement. Considering this, we attempted to develop EST-derived-SSR (eSSR) markers and utilize them in germplasm characterization, cross-genera transferability and in silico comparative mapping. From 66,027 foxtail millet EST sequences 24,828 non-redundant ESTs were deduced, representing ~16 Mb, which revealed 534 (~2%) eSSRs in 495 SSR containing ESTs at a frequency of 1/30 kb. A total of 447 pp were successfully designed, of which 327 were mapped physically onto nine chromosomes. About 106 selected primer pairs representing the foxtail millet genome showed high-level of cross-genera amplification at an average of ~88% in eight millets and four non-millet species. Broad range of genetic diversity (0.02-0.65) obtained in constructed phylogenetic tree using 40 eSSR markers demonstrated its utility in germplasm characterizations and phylogenetics. Comparative mapping of physically mapped eSSR markers showed considerable proportion of sequence-based orthology and syntenic relationship between foxtail millet chromosomes and sorghum (~68%), maize (~61%) and rice (~42%) chromosomes. Synteny analysis of eSSRs of foxtail millet, rice, maize and sorghum suggested the nested chromosome fusion frequently observed in grass genomes. Thus, for the first time we had generated large-scale eSSR markers in foxtail millet and demonstrated their utility in germplasm characterization, transferability, phylogenetics and comparative mapping studies in millets and bioenergy grass species.Item Development and utilization of novel SSRs in foxtail millet [Setaria italica (L.) P. Beauv.](John Wiley & Sons, 2013) Gupta, Sarika; Kumari, Kajal; Muthamilarasan, Mehanathan; Subramanian, Alagesan; Prasad, ManojAlthough the foxtail millet [Setaria italica (L.) P. Beauv.] is recently regarded as a model crop for studying functional genomics of biofuel grasses, its genetic improvement to some extent was limited due to the non-availability of molecular markers, particularly the microsatellite markers and the saturated genetic linkage map. Considering this, we attempted to generate a significant number of microsatellite markers in cultivar ‘Prasad’. Two hundred and fifty-six clones were sequenced to generate 41.82-kb high-quality sequences retrieved from genomic library enriched with dinucleotide repeat motifs. Microsatellites were identified in 194 (76%) of the 256 positive clones, and 64 primer pairs (pp) were successfully designed from 95 (49%) unique SSR-containing clones. The 67.4% primer designing ability, 100% PCR amplification efficiency and 45.3% polymorphic potential in the parents of F2 mapping population established the efficacy of genomic microsatellites. All the 64 microsatellite markers displayed high level of cross-species amplification (~67%) in 10 millets and non-millets species. These experimental findings suggest the utility and efficacy of SSRs in diverse genotyping applications, resolving QTLs, phylogenetic relationships and transferability in several important grass species.Item Transcriptome analysis of differentially expressed genes during embryosac development in apomeiotic non parthenogenetic interspecific hybrid of Pennisetum glaucum(Springer, 2012) Sahu, Pranav Pankaj; Gupta, Sarika; Malaviya, D. R.; Roy, Ajoy Kumar; Kaushal, Pankaj; Prasad, ManojApomixis results in the production of genetically uniform progeny, derived from the fertilization independent development (parthenogenesis) of an unreduced egg cell (apomeiosis). To identify genes involved in the apomeiosis, a comparative transcriptome analysis of differentially expressed genes during embryo sac (ES) development in a sexual Pennisetum glaucum (genotype 81A1) and its apomeiotic (aposporic) non-parthenogenetic interspecific hybrid (BC1GO) was investigated. BC1GO exhibited the partitioned apomeiosis component, whereby the second apomixis component viz., parthenogenesis was completely lacking. A total of 96 non-redundant transcripts were recovered using suppression subtractive hybridization and classified into 11 different categories according to their putative functions. Amongst the identified transcripts, many of them belonged to unknown function (40%) followed by those involved in protein metabolism, stress response, pollen/ovule/embryo development, and translation/protein modification process. A data search of transcriptional profiling in other apomictic species revealed that 75% of the differentially expressed transcripts have not been reported in previous studies. By macroarray analysis, we identified differential expression pattern of 96 transcripts, 45 (47%) of which showed ≥2-fold induction in apomeiotic BC1GO. Further, the obtained results were validated by quantitative real-time polymerase chain reaction to have a comparative expression profiling of eight selected up-regulated transcripts (≥2.5-fold) between BC1GO and 81A1 at different phases of ovule development. In silico mapping demonstrated that 13 transcripts were located onto rice chromosome 2, region syntenic with the apospory locus as reported in Brachiaria brizantha and Paspalum notatum. The expression patterns of these transcripts showed a significant difference at differentiating megaspore mother cell and gametogenesis stages thereby suggesting their involvement in floral development during apomeiotic (Panicum-type aposporous) ES development.Item Sequence based novel genomic microsatellite markers for robust genotyping purposes in foxtail millet [Setaria italica (L.) P. Beauv.](Springer Science, 2012) Gupta, Sarika; Kumari, Kajal; Sahu, Pranav Pankaj; Vidapu, Sudhakar; Prasad, ManojThe unavailability of microsatellite markers and saturated genetic linkage map has restricted the genetic improvement of foxtail millet [Setaria italica (L.) P. Beauv.], despite the fact that in recent times it has been documented as a new model species for biofuel grasses. With the objective to generate a good number of microsatellite markers in foxtail millet cultivar 'Prasad', 690 clones were sequenced which generated 112.95 kb high quality sequences obtained from three genomic libraries each enriched with different microsatellite repeat motifs. Microsatellites were identified in 512 (74.2%) of the 690 positive clones and 172 primer pairs (pp) were successfully designed from 249 (48.6%) unique SSR-containing clones. The efficacies of the microsatellite containing genomic sequences were established by superior primer designing ability (69%), PCR amplification efficiency (85.5%) and polymorphic potential (52%) in the parents of F(2) mapping population. Out of 172 pp, functional 147 markers showed high level of cross-species amplification (~74%) in six grass species. Higher polymorphism rate and broad range of genetic diversity (0.30-0.69 averaging 0.58) obtained in constructed phylogenetic tree using 52 microsatellite markers, demonstrated the utility of markers in germplasm characterizations. In silico comparative mapping of 147 foxtail millet microsatellite containing sequences against the mapping data of sorghum (~18%), maize (~16%) and rice (~5%) indicated the presence of orthologous sequences of the foxtail millet in the respective species. The result thus demonstrates the applicability of microsatellite markers in various genotyping applications, determining phylogenetic relationships and comparative mapping in several important grass species.Item Molecular marker technology in plants(IK International Publishers, 2012) Gupta, Sarika; Ghose, Tapas Kumar; Jaiswal, J. P.; Prasad, Manoj