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Author: search.filters.author.Tyagi, Akhilesh K.Subject: search.filters.subject.Transcriptome

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    Cytological, transcriptome and miRNome temporal landscapes decode enhancement of rice grain size
    (BioMed Central Ltd, 2023) Mahto, Arunima; Yadav, Antima; Aswathi, P. V.; Parida, Swarup K.; Tyagi, Akhilesh K.; Agarwal, Pinky
    Background Rice grain size (GS) is an essential agronomic trait. Though several genes and miRNA modules influencing GS are known and seed development transcriptomes analyzed, a comprehensive compendium connecting all possible players is lacking. This study utilizes two contrasting GS indica rice genotypes (small-grained SN and large-grained LGR). Rice seed development involves five stages (S1–S5). Comparative transcriptome and miRNome atlases, substantiated with morphological and cytological studies, from S1–S5 stages and flag leaf have been analyzed to identify GS proponents. Results Histology shows prolonged endosperm development and cell enlargement in LGR. Stand-alone and comparative RNAseq analyses manifest S3 (5–10 days after pollination) stage as crucial for GS enhancement, coherently with cell cycle, endoreduplication, and programmed cell death participating genes. Seed storage protein and carbohydrate accumulation, cytologically and by RNAseq, is shown to be delayed in LGR. Fourteen transcription factor families influence GS. Pathway genes for four phytohormones display opposite patterns of higher expression. A total of 186 genes generated from the transcriptome analyses are located within GS trait-related QTLs deciphered by a cross between SN and LGR. Fourteen miRNA families express specifically in SN or LGR seeds. Eight miRNA-target modules display contrasting expressions amongst SN and LGR, while 26 (SN) and 43 (LGR) modules are differentially expressed in all stages. Conclusions Integration of all analyses concludes in a “Domino effect” model for GS regulation highlighting chronology and fruition of each event. This study delineates the essence of GS regulation, providing scope for future exploits. The rice grain development database (RGDD) ( www.nipgr.ac.in/RGDD/index.php; https://doi.org/10.5281/zenodo.7762870) has been developed for easy access of data generated in this paper.
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    Expanding frontiers in plant transcriptomics in aid of functional genomics and molecular breeding
    (John Wiley & Sons Ltd, 2014) Agarwal, Pinky; Parida, Swarup K.; Mahto, Arunima; Das, Sweta; Mathew, Iny Elizebeth; Malik, Naveen; Tyagi, Akhilesh K.
    The transcript pool of a plant part, under any given condition, is a collection of mRNAs that will pave the way for a biochemical reaction of the plant to stimuli. Over the past decades, transcriptome study has advanced from Northern blotting to RNA sequencing (RNA-seq), through other techniques, of which real-time quantitative polymerase chain reaction (PCR) and microarray are the most significant ones. The questions being addressed by such studies have also matured from a solitary process to expression atlas and marker-assisted genetic enhancement. Not only genes and their networks involved in various developmental processes of plant parts have been elucidated, but also stress tolerant genes have been highlighted. The transcriptome of a plant with altered expression of a target gene has given information about the downstream genes. Marker information has been used for breeding improved varieties. Fortunately, the data generated by transcriptome analysis has been made freely available for ample utilization and comparison. The review discusses this wide variety of transcriptome data being generated in plants, which includes developmental stages, abiotic and biotic stress, effect of altered gene expression, as well as comparative transcriptomics, with a special emphasis on microarray and RNA-seq. Such data can be used to determine the regulatory gene networks, which can subsequently be utilized for generating improved plant varieties.
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    Transcriptome analysis of rin mutant fruit and in silico analysis of promoters of differentially regulated genes provides insight into LeMADS-RIN-regulated ethylene-dependent as well as ethylene-independent aspects of ripening in tomato
    (Springer Science, 2012) Kumar, Rahul; Sharma, Manoj K.; Kapoor, Sanjay; Tyagi, Akhilesh K.; Sharma, Arun K.
    A thorough understanding of molecular mechanisms underlying ripening is the prerequisite for genetic manipulation of fruits for better shelf-life and nutritional quality. Mutation in LeMADS-RIN, a MADS-box gene, leads to non-ripening phenotype of rin fruits in tomato. Characterization of ripening-inhibitor (rin) mutant has elucidated important role of ethylene in the regulation of climacteric fruit ripening. A complete understanding of this mutation will unravel novel genetic regulatory mechanisms involved in fruit ripening. In this study, fruit transcriptomes of two genotypes, including a cultivated Indian cultivar Solanum lycopersicum cv. Pusa Ruby and a homozygous line harboring the rin mutation (LA1795) were compared to get better insight into RIN-regulated ethylene-dependent and ethylene-independent events during ripening. Cluster analysis of ripening-related genes indicated a major shift in their expression profiles in rin mutant fruit. A total of 112 genes, exhibiting expression patterns similar to that of LeMADS-RIN in wild-type fruits, showed down regulation of expression in the rin mutant. In silico analysis of putative promoters of these genes for the presence of CArG box along with ERE and ethylene inducibility of these genes revealed that genes lacking CArG box in their regulatory regions could be indirectly regulated by LeMADS-RIN. New regulators of ethylene-dependent aspect of ripening were also identified. In this study, we have made an attempt to distinguish between ethylene-dependent and ethylene-independent aspects of ripening, which will be useful for developing strategies to improve fruit-related agronomic traits in tomato and other crops.
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    Comparative analysis of kabuli chickpea transcriptome with desi and wild chickpea provides a rich resource for development of functional markers
    (Public Library of Science, 2012) Agarwal, Gaurav; Jhanwar, Shalu; Priya, Pushp; Singh, Vikash K.; Saxena, Maneesha S.; Parida, Swarup K.; Garg, Rohini; Tyagi, Akhilesh K.; Jain, Mukesh
    Chickpea (Cicer arietinum L.) is an important crop legume plant with high nutritional value. The transcriptomes of desi and wild chickpea have already been sequenced. In this study, we sequenced the transcriptome of kabuli chickpea, C. arietinum (genotype ICCV2), having higher commercial value, using GS-FLX Roche 454 and Illumina technologies. The assemblies of both Roche 454 and Illumina datasets were optimized using various assembly programs and parameters. The final optimized hybrid assembly generated 43,389 transcripts with an average length of 1065 bp and N50 length of 1653 bp representing 46.2 Mb of kabuli chickpea transcriptome. We identified a total of 5409 simple sequence repeats (SSRs) in these transcript sequences. Among these, at least 130 and 493 SSRs were polymorphic with desi (ICC4958) and wild (PI489777) chickpea, respectively. In addition, a total of 1986 and 37,954 single nucleotide polymorphisms (SNPs) were predicted in kabuli/desi and kabuli/wild genotypes, respectively. The SNP frequency was 0.043 SNP per kb for kabuli/desi and 0.821 SNP per kb for kabuli/wild, reflecting very low genetic diversity in chickpea. Further, SSRs and SNPs present in tissue-specific and transcription factor encoding transcripts have been identified. The experimental validation of a selected set of polymorphic SSRs and SNPs exhibited high intra-specific polymorphism potential between desi and kabuli chickpea, suggesting their utility in large-scale genotyping applications. The kabuli chickpea gene index assembled, and SSRs and SNPs identified in this study will serve as useful genomic resource for genetic improvement of chickpea.
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    Identification and characterization of high temperature stress responsive genes in bread wheat (Triticum aestivum L.) and their regulation at various stages of development
    (Springer Science, 2011) Chauhan, Harsh; Khurana, Neetika; Tyagi, Akhilesh K.; Khurana, Jitendra P.; Khurana, Paramjit
    To elucidate the effect of high temperature, wheat plants (Triticum aestivum cv. CPAN 1676) were given heat shock at 37 and 42°C for 2 h, and responsive genes were identified through PCR-Select Subtraction technology. Four subtractive cDNA libraries, including three forward and one reverse subtraction, were constructed from three different developmental stages. A total of 5,500 ESTs were generated and 3,516 high quality ESTs submitted to Genbank. More than one-third of the ESTs generated fall in unknown/no hit category upon homology search through BLAST analysis. Differential expression was confirmed by cDNA macroarray and by northern/RT-PCR analysis. Expression analysis of wheat plants subjected to high temperature stress, after 1 and 4 days of recovery, showed fast recovery in seedling tissue. However, even after 4 days, recovery was negligible in the developing seed tissue after 2 h of heat stress. Ten selected genes were analyzed in further detail including one unknown protein and a new heat shock factor, by quantitative real-time PCR in an array of 35 different wheat tissues representing major developmental stages as well as different abiotic stresses. Tissue specificity was examined along with cross talk with other abiotic stresses and putative signalling molecules.