32 research outputs found
Genotyping-by-sequencing based intra-specific genetic map refines a ‘‘QTL-hotspot” region for drought tolerance in chickpea
No full textTo enhance the marker density in the “QTL-hotspot” region, harboring several QTLs for drought tolerance-related traits identified on linkage group 04 (CaLG04) in chickpea recombinant inbred line (RIL) mapping population ICC 4958 × ICC 1882, a genotyping-by-sequencing approach was adopted. In total, 6.24 Gb data from ICC 4958, 5.65 Gb data from ICC 1882 and 59.03 Gb data from RILs were generated, which identified 828 novel single-nucleotide polymorphisms (SNPs) for genetic mapping. Together with these new markers, a high-density intra-specific genetic map was developed that comprised 1,007 marker loci spanning a distance of 727.29 cM. QTL analysis using the extended genetic map along with precise phenotyping data for 20 traits collected over one to seven seasons identified 49 SNP markers in the “QTL-hotspot” region. These efforts have refined the “QTL-hotspot” region to 14 cM. In total, 164 main-effect QTLs including 24 novel QTLs were identified. In addition, 49 SNPs integrated in the “QTL-hotspot” region were converted into cleaved amplified polymorphic sequence (CAPS) and derived CAPS (dCAPS) markers which can be used in marker-assisted breeding
Prioritization of candidate genes in "QTL-hotspot" region for drought tolerance in chickpea (Cicer arietinum L.)
No full textA combination of two approaches, namely QTL analysis and gene enrichment analysis were used to identify candidate genes in the "QTL-hotspot" region for drought tolerance present on the Ca4 pseudomolecule in chickpea. In the first approach, a high-density bin map was developed using 53,223 single nucleotide polymorphisms (SNPs) identified in the recombinant inbred line (RIL) population of ICC 4958 (drought tolerant) and ICC 1882 (drought sensitive) cross. QTL analysis using recombination bins as markers along with the phenotyping data for 17 drought tolerance related traits obtained over 1-5 seasons and 1-5 locations split the "QTL-hotspot" region into two subregions namely "QTL-hotspot_a" (15 genes) and "QTL-hotspot_b" (11 genes). In the second approach, gene enrichment analysis using significant marker trait associations based on SNPs from the Ca4 pseudomolecule with the above mentioned phenotyping data, and the candidate genes from the refined "QTL-hotspot" region showed enrichment for 23 genes. Twelve genes were found common in both approaches. Functional validation using quantitative real-time PCR (qRT-PCR) indicated four promising candidate genes having functional implications on the effect of "QTL-hotspot" for drought tolerance in chickpea.Sandip M Kale, Deepa Jaganathan, Pradeep Ruperao, Charles Chen, Ramu Punna, Himabindu Kudapa, Mahendar Thudi, Manish Roorkiwal, Mohan AVSK Katta, Dadakhalandar Doddamani, Vanika Garg, P B Kavi Kishor, Pooran M Gaur, Henry T Nguyen, Jacqueline Batley, David Edwards, Tim Sutton and Rajeev K Varshne
Genotyping-by-sequencing based intra-specific genetic map refines a ‘‘QTL-hotspot” region for drought tolerance in chickpea
No full textTo enhance the marker density in the "QTL-hotspot" region, harboring several QTLs for drought tolerance-related traits identified on linkage group 04 (CaLG04) in chickpea recombinant inbred line (RIL) mapping population ICC 4958 × ICC 1882, a genotyping-by-sequencing approach was adopted. In total, 6.24 Gb data from ICC 4958, 5.65 Gb data from ICC 1882 and 59.03 Gb data from RILs were generated, which identified 828 novel single-nucleotide polymorphisms (SNPs) for genetic mapping. Together with these new markers, a high-density intra-specific genetic map was developed that comprised 1,007 marker loci spanning a distance of 727.29 cM. QTL analysis using the extended genetic map along with precise phenotyping data for 20 traits collected over one to seven seasons identified 49 SNP markers in the "QTL-hotspot" region. These efforts have refined the "QTL-hotspot" region to 14 cM. In total, 164 main-effect QTLs including 24 novel QTLs were identified. In addition, 49 SNPs integrated in the "QTL-hotspot" region were converted into cleaved amplified polymorphic sequence (CAPS) and derived CAPS (dCAPS) markers which can be used in marker-assisted breeding.Deepa Jaganathan, Mahendar Thudi, Sandip Kale, Sarwar Azam, Manish Roorkiwal, Pooran M. Gaur, P B Kavi Kishor, Henry Nguyen, Tim Sutton, Rajeev K. Varshne
CRISPR for Crop Improvement: An Update Review
No full textThe availability of genome sequences for several crops and advances in genome editing approaches has opened up possibilities to breed for almost any given desirable trait. Advancements in genome editing technologies such as zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) has made it possible for molecular biologists to more precisely target any gene of interest. However, these methodologies are expensive and time-consuming as they involve complicated steps that require protein engineering. Unlike first-generation genome editing tools, CRISPR/Cas9 genome editing involves simple designing and cloning methods, with the same Cas9 being potentially available for use with different guide RNAs targeting multiple sites in the genome. After proof-of-concept demonstrations in crop plants involving the primary CRISPR-Cas9 module, several modified Cas9 cassettes have been utilized in crop plants for improving target specificity and reducing off-target cleavage (e.g., Nmcas9, Sacas9, and Stcas9). Further, the availability of Cas9 enzymes from additional bacterial species has made available options to enhance specificity and efficiency of gene editing methodologies. This review summarizes the options available to plant biotechnologists to bring about crop improvement using CRISPR/Cas9 based genome editing tools and also presents studies where CRISPR/Cas9 has been used for enhancing biotic and abiotic stress tolerance. Application of these techniques will result in the development of non-genetically modified (Non-GMO) crops with the desired trait that can contribute to increased yield potential under biotic and abiotic stress conditions
Fine mapping and gene cloning in the post-NGS era: advances and prospects
No full textImprovement in traits of agronomic importance is the top breeding priority of crop improvement programs. Majority of
these agronomic traits show complex quantitative inheritance. Identification of quantitative trait loci (QTLs) followed by
fine mapping QTLs and cloning of candidate genes/QTLs is central to trait analysis. Advances in genomic technologies
revolutionized our understanding of genetics of complex traits, and genomic regions associated with traits were employed in
marker-assisted breeding or cloning of QTLs/genes. Next-generation sequencing (NGS) technologies have enabled genomewide
methodologies for the development of ultra-high-density genetic linkage maps in different crops, thus allowing placement
of candidate loci within few kbs in genomes. In this review, we compare the marker systems used for fine mapping and
QTL cloning in the pre- and post-NGS era. We then discuss how different NGS platforms in combination with advanced
experimental designs have improved trait analysis and fine mapping. We opine that efficient genotyping/sequencing assays
may circumvent the need for cumbersome procedures that were earlier used for fine mapping. A deeper understanding of
the trait architectures of agricultural significance will be crucial to accelerate crop improvement
Prioritization of candidate genes in “QTL-hotspot” region for drought tolerance in chickpea (Cicer arietinum L.)
No full textA combination of two approaches, namely QTL analysis and gene enrichment analysis were used
to identify candidate genes in the “QTL-hotspot” region for drought tolerance present on the Ca4
pseudomolecule in chickpea. In the first approach, a high-density bin map was developed using
53,223 single nucleotide polymorphisms (SNPs) identified in the recombinant inbred line (RIL)
population of ICC 4958 (drought tolerant) and ICC 1882 (drought sensitive) cross. QTL analysis using
recombination bins as markers along with the phenotyping data for 17 drought tolerance related
traits obtained over 1–5 seasons and 1–5 locations split the “QTL-hotspot” region into two subregions
namely “QTL-hotspot_a” (15 genes) and “QTL-hotspot_b” (11 genes). In the second approach,
gene enrichment analysis using significant marker trait associations based on SNPs from the Ca4
pseudomolecule with the above mentioned phenotyping data, and the candidate genes from the
refined “QTL-hotspot” region showed enrichment for 23 genes. Twelve genes were found common
in both approaches. Functional validation using quantitative real-time PCR (qRT-PCR) indicated four
promising candidate genes having functional implications on the effect of “QTL-hotspot” for drought
tolerance in chickpea
Morphophysiological dormancy and germination ecology in diaspores of the subtropical palm Phoenix canariensis Chabaud
No full textThe timing of germination is a crucial event in a plant’s life cycle. Seed dormancy and germination mechanisms are important factors regulating seedling emergence. Since detailed experimental evidence for germination pattern of Phoenix canariensis colonizing sub-tropical climate is scarce, we investigated seed dormancy and germination ecology of P. canariensis. We found that the embryo is underdeveloped at the time of dispersal and doubles in size before the cotyledonary petiole (CP) protrudes through the operculum. The primary root and plumule emerge from the elongated CP outside the seed. In light/dark at 30/25C, the CP emerged from 8% of the diaspores within 30 days and from 76% within 14 weeks. Thus, 8% of the diaspores have MD and the others MPD. Removal of the pericarp and operculum resulted in 100% germination within 5 days in light/dark at 30/25C. Cold and warm stratification as well as treatment with GA3 significantly increased the germination speed, but the final germination percentage was not significantly increased. Seed germination was synchronized in early summer when seed dormancy was released by cold stratification in the soil over winter. A remote-tubular germination type and intricate root system provide an ecological advantage to the seedling establishment.The presentation of the authors' names and (or) special characters in the title of the pdf file of the accepted manuscript may differ slightly from what is displayed on the item page. The information in the pdf file of the accepted manuscript reflects the original submission by the author
Sentrin/SUMO Specific Proteases as Novel Tissue-Selective Modulators of Vitamin D Receptor-Mediated Signaling
No full textabstract: Vitamin D receptor (VDR) is a substrate for modification with small ubiquitin-like modifier (SUMO). To further assess the role of reversible SUMOylation within the vitamin D hormonal response, we evaluated the effects of sentrin/SUMO-specific proteases (SENPs) that can function to remove small ubiquitin-like modifier (SUMO) from target proteins upon the activities of VDR and related receptors. We report that SENP1 and SENP2 strikingly potentiate ligand-mediated transactivation of VDR and also its heterodimeric partner, retinoid X receptor (RXRα) with depletion of cellular SENP1 significantly diminishing the hormonal responsiveness of the endogenous vitamin D target gene CYP24A1. We find that SENP-directed modulation of VDR activity is cell line-dependent, achieving potent modulatory effects in Caco-2 and HEK-293 cells, while in MCF-7 cells the vitamin D signal is unaffected by any tested SENP. In support of their function as novel modulators of the vitamin D hormonal pathway we demonstrate that both SENP1 and SENP2 can interact with VDR and reverse its modification with SUMO2. In a preliminary analysis we identify lysine 91, a residue known to be critical for formation and DNA binding of the VDR-RXR heterodimer, as a minor SUMO acceptor site within VDR. In combination, our results support a repressor function for SUMOylation of VDR and reveal SENPs as a novel class of VDR/RXR co-regulatory protein that significantly modulate the vitamin D response and which could also have important impact upon the functionality of both RXR-containing homo and heterodimers.The article is published at http://journals.plos.org/plosone/article?id=10.1371/journal.pone.008950
Creation of novel alleles of fragrance gene OsBADH2 in rice through CRISPR/Cas9 mediated gene editing.
No full textFragrance in rice grains is a key quality trait determining its acceptability and marketability. Intensive research on rice aroma identified mutations in betaine aldehyde dehydrogenase (OsBADH2) leading to production of aroma in rice. Gene editing technologies like CRISPR/Cas9 system has opened new avenues for accelerated improvement of rice grain quality through targeted mutagenesis. In this study, we have employed CRISPR/Cas9 tool to create novel alleles of OsBADH2 leading to introduction of aroma into an elite non-aromatic rice variety ASD16. PCR analysis of putative transformants using primers targeting the flanking regions of sgRNA in the 7th exon of OsBADH2 identified 37.5% potential multi-allelic mutations in T0 generation. Sensory evaluation test in the leaves of T0 lines identified thirteen lines belonging to five independent events producing aroma. Sequence analysis of these aromatic T0 lines identified 22 different types of mutations located within -17 bp to +15bp of sgRNA region. The -1/-2 bp deletion in the line # 8-19 and -8/-5 bp deletion in the line # 2-16 produced strong aroma and the phenotype was stably inherited in the T1 generation. Comparative volatile profiling detected novel aromatic compounds viz., pyrrolidine, pyridine, pyrazine, pyradazine and pyrozole in the grains of T1 progenies of line # 8-19. This study has demonstrated the use of CRISPR/Cas9 in creating novel alleles of OsBADH2 to introduce aroma into any non-aromatic rice varieties
