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20211
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Subject: Agricultural Biotechnology × Author: Bhatt, Megha × End date: 2024 × Start date: 2020 × Type: Thesis ×
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    ThesisItemOpen Access
    Isolation, cloning and in silico analysis of drought responsive EcMyb1 transcription factor gene from Eleusine coracana
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-01) Bhatt, Megha; Lohani, Pushpa
    Myb genes are members of a family of transcription factors that play an important role in cell division, differentiation, response to abiotic stress etc. It has been reported that MYB transcription factor play an important role as a master regulator and modulates the expression of various downstream genes involved in drought stress response. The partial sequence of Ecmyb1 gene was isolated and submitted to NCBI with accession number JN107890.1. In present study, full length Ecmyb1 gene was isolated and cloned. The cloned fragment was sequenced and submitted to NCBI with accession number MT312253. For expression analysis of Ecmyb1 gene, finger millet drought tolerant variety PRM 6107 was given three different abiotic stress treatment i.e. drought, salt and heat stress. The expression of the gene increased from 12 to 28 fold under drought stress as compared to control. Similarly, in salt and heat treated plants, expression increased from 7 to 29 fold and 19 to 46 fold respectively. The cloned gene was introgressed into Arabidopsis thaliana plant by floral dip method. The transformed plants were selected by hygromysin screening and confirmed by PCR using hygromycin primers. Insilco analysis of Ecmyb1 gene was carried out using various softwares. Promoter analysis of Ecmyb1 gene revealed the presence of various regulatory elements like ABA responsive elements (CACT GG), stress responsive elements (AAGG GG), Myb recognition sites (CAACAG), Myb binding sites (CAAC TG), CpG island and tandem repeats. Homology and phylogenetic analysis of Ecmyb1 gene and amino acid sequences showed similarity with other MYB proteins such as Sateria italica, Hordeum vulgare, Saccharum barberi and Oryza sativa etc. Motif analysis predicted the presence of HTH domain in EcMYB1 protein. Subsequently, it was computed that EcMYB1 protein is water soluble, thermostable and slightly acidic in nature. Additionally, it was observed that EcMYB1 protein contains 28% alpha helix, 5.43% extended strand, 2.57% beta turn and 64% random coil. 3-D structure of EcMYB1 using swiss model predicted that 90% aminoacid residues were in Ramachandran favored region with no bad angles. It indicated that EcMYB1 protein may exist in nature.