RAPID AND EFFICIENT HIGH QUALITY VIRAL RNA ISOLATION FROM BANANA BRACT MOSAIC VIRUS INFECTED BANANA CV. GRAND NAINE

S.C. EKATPURE¹*, K.B. SONI², P.R. JADHAV³, R.S. LEKSHMI⁴, S. ALEX^5
1Department of Plant Biotechnology, College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala,695522, India
²Department of Plant Biotechnology, College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala,695522, India
³Department of Plant Biotechnology, College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala,695522, India
⁴Department of Plant Biotechnology, College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala,695522, India
⁵Department of Plant Biotechnology, College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala,695522, India
* Corresponding Author : sachi.ekatpure@gmail.com

Received : 15-03-2019     Accepted : 26-03-2019     Published : 30-03-2019
Volume : 11     Issue : 6       Pages : 8096 - 8099
Int J Agr Sci 11.6 (2019):8096-8099

Keywords : Banana Bract Mosaic Virus (BBrMV), ssRNA viruses, RNA isolation, Replicase gene, Grand Naine, Polysaccharides and Phenolics
Conflict of Interest : None declared
Acknowledgements/Funding : Authors are thankful to the Kerala State Council for Science and Technology (KSCSTE), Government of Kerala and College of Agriculture, Kerala Agricultural University, Vellayani, Thiruvananthapuram, Kerala, 695522, India
Author Contribution : All authors equally contributed

Isolation of high quality viral RNA from a banana plant is quite difficult, due to the presence of high amount of polysaccharides and phenolic compounds in the plant. These compounds not only reduce the yield of RNA but also make it unsuitable for the molecular studies, like cDNA library construction, molecular analysis and RNAseq. In the present study we describe a simple RNA isolation procedure based on Rodriguez-Garcia protocol to isolate high-quality viral RNA from banana plant infected with Banana Bract Mosaic Virus (BBrMV). This modified protocol effectively removes phenols and polysaccharides and gives a yield of 100-200 μg high-quality total RNA per gram of fresh tissue. PCR product of partial BBrMV replicase and movement protein gene from bract sample was sequenced generating 386 nt and 339 nt sequences respectively which were submitted to NCBI database. These sequences showed similarity to replicase (96 %) and movement protein gene (97 %) to that of Trichy isolate, with 100 percent query cover in BLAST result. Isolated RNA showed purity suitable for northern analysis, RT-PCR and cDNA library construction.

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