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Identification of Heat-Related ESTs in Moth Bean Through Suppression Subtraction Hybridization

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Abstract

Moth bean (Vigna aconitifolia (Jacq.) Marechal), an important grain-legume crop grown in hot desert regions of Thar, under scorching sun rays, was investigated for heat tolerance at molecular level. In the present study, we constructed a forward suppression subtractive hybridization (SSH) cDNA library of heat tolerant genotype RMO-40 to identify genes expressing under delayed response to elevated temperature. Heat induction was carried out by exposing 14-day-old seedlings to elevated temperature of 42 °C for 30 min. A total of 125 unigenes (33 contigs and 92 singletons) were derived by cluster assembly and sequence alignment of 200 ESTs; out of 125 unigenes, 21 (16 %) were found to be novel to moth bean. Gene ontology functional classification terms were retrieved for 98 (78.4 %) unigenes of which 73 (58.4 %) ESTs were functionally annotated (GO consensus) where 19 unigenes were annotated with 11 enzyme commission (EC) codes and were mapped to 25 different KEGG pathways. We have identified a majority of heat-shock proteins (constituting 35 % of the present library) aiding heat stress tolerance to moth bean. An expression level of 22 ESTs generated from the above SSH cDNA library was studied through semiquantitative RT-PCR assay simultaneously under 5 and 30 min of heat stress at 42 °C.

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Authors and Affiliations

  1. Plant Biotechnology Centre, Swami Keshwan and Rajasthan Agricultural University, Bikaner, Rajasthan, 334006, India

    Karun Gurjar, Sakshi Rampuria, Uma Joshi & Ramavtar Sharma

  2. National Research Centre on Plant Biotechnology, IARI Campus, New Delhi, 110012, India

    Paramita Palit & T. Mohapatra

  3. National Bureau of Plant Genetic Resources, IARI Campus, New Delhi, 110012, India

    K. V. Bhatt

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  1. Karun Gurjar
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  2. Sakshi Rampuria
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  3. Uma Joshi
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  4. Paramita Palit
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  5. K. V. Bhatt
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  6. T. Mohapatra
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  7. Ramavtar Sharma
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Correspondence to Ramavtar Sharma.

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Gurjar, K., Rampuria, S., Joshi, U. et al. Identification of Heat-Related ESTs in Moth Bean Through Suppression Subtraction Hybridization. Appl Biochem Biotechnol 173, 2116–2128 (2014). https://doi.org/10.1007/s12010-014-1011-2

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  • DOI: https://doi.org/10.1007/s12010-014-1011-2

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