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miR430: the novel heat-responsive microRNA identified from miRNome analysis in wheat (Triticum aestivum L.)

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Abstract

miRNAs are 21–24 nt non-coding RNAs involved in negative regulation of the target gene expression in response to stress and developmental cues. Wheat, the major staple food crop, is highly sensitive to heat stress (HS). Even moderate high temperatures during critical stages causes reduction in yield and quality of the grains. Here, we identified a novel miRNA (candidate miR430) from wheat using de novo assembly, and cloned it from wheat cv. HD2985 using universal adaptor. The identified miRNA was mapped on to the Chromosome 3B and predicted to have more than 30 target genes. Most of the targets identified were associated with heat shock proteins. Expression analysis of miR430 in contrasting wheat cultivars by quantitative real-time PCR revealed significant genotypic variations under HS. Tissue specific expression analysis showed relatively low expression in leaves, as compared with stem and root under HS. A negative correlation was found between the expression of miR430 and their respective target genes under HS. MiR430 can be used to manipulate the expression of target genes under HS towards enhancing HS tolerance for the development of ‘climate-smart’ wheat crop.

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Acknowledgments

The financial support received from Indian Council of Agriculture Research (ICAR) under the National Innovations for Climate Resilient Agriculture (NICRA) Project (12/115 TG3079) and extra-mural research funding by SERB, Department of Science and Technology (DST) (Project no. SERB/SB/SO/PS/07/2014) is highly acknowledged.

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  1. Mahesh Kumar and Ranjeet R. Kumar have contributed equally to this work.

Authors and Affiliations

  1. Division of Biochemistry, ICAR-Indian Agricultural Research Institute, New Delhi, 110012, India

    Mahesh Kumar, Ranjeet R. Kumar, Suneha Goswami, Pooja Verma, Raj D. Rai & Shelly Praveen

  2. Division of Plant Physiology, ICAR-Indian Agricultural Research Institute, New Delhi, 110012, India

    Viswanathan Chinnusamy

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  1. Mahesh Kumar
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Contributions

MK, RRK—conceived and designed the experiments. MK, SG, RRK—involved in isolation and cloning of miRNA. PV, MK—performed transcript profiling. RRK, RDR, VC, SP—wrote the paper and edited the manuscript.

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Correspondence to Ranjeet R. Kumar.

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The authors declare that they have no competing interest.

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40502_2017_341_MOESM1_ESM.docx

Fig S1. Total microRNA isolated from different tissues of wheat cultivars HD2985 (thermotolerant) and HD2329 (thermosusceptible) under control (22 °C) and heat stress (42 °C, 2 h) treatments; miPremier microRNA isolation kit (Sigma-Aldrich, UK) was used for the isolation; 12% gel was used for the SDS-PAGE; M1- 100 bp DNA ladder, M2- 50 bp DNA ladder. Fig S2. PCR amplification and restriction analysis of novel microRNA identified based on miRNome analysis of wheat, (a) Amplicon of miR430, (b) Restriction analysis of plasmid with EcoRI; mature miR430 specific forward primer was used along with universal reverse primer (as given in the kit) for the amplification, amplicon was cloned in pGEMTEasy vector (Promega, UK) before sequencing by Sangers di-deoxy method, 3% agarose gel was used for the visualization. Fig S3. Pre-mature sequence of novel miR430 amplified from HD2985 cultivar of wheat; Sangers di-deoxy method was used for the sequencing, shaded sequence shows the mature sequence of the identified miRNA. (DOCX 896 kb)

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Kumar, M., Kumar, R.R., Goswami, S. et al. miR430: the novel heat-responsive microRNA identified from miRNome analysis in wheat (Triticum aestivum L.). Ind J Plant Physiol. 22, 566–576 (2017). https://doi.org/10.1007/s40502-017-0341-9

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