African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12191

Full Length Research Paper

Modified protocol for RNA isolation from different parts of field-grown jute plant suitable for NGS data generation and quantitative real-time RT-PCR

Rasel Ahmed
  • Rasel Ahmed
  • Basic and Applied Research on Jute Project, Bangladesh Jute Research Institute, Dhaka, Bangladesh.
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Md. Sabbir Hossain
  • Md. Sabbir Hossain
  • Basic and Applied Research on Jute Project, Bangladesh Jute Research Institute, Dhaka, Bangladesh.
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Md. Samiul Haque
  • Md. Samiul Haque
  • Basic and Applied Research on Jute Project, Bangladesh Jute Research Institute, Dhaka, Bangladesh.
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Md. Monjurul Alam
  • Md. Monjurul Alam
  • Basic and Applied Research on Jute Project, Bangladesh Jute Research Institute, Dhaka, Bangladesh.
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Md. Shahidul Islam
  • Md. Shahidul Islam
  • Basic and Applied Research on Jute Project, Bangladesh Jute Research Institute, Dhaka, Bangladesh.
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  •  Received: 31 March 2019
  •  Accepted: 17 June 2019
  •  Published: 31 July 2019

Abstract

High quality RNA extraction from field-grown jute plants can be difficult due to the presence of complex polyphenolic, polysaccharide and waxes. But isolation of RNA in high quality is a basic need in plant genetics, genomics, transcriptomics, molecular biology and related physiological investigations. Here, cetyl trimethyl ammonium bromide (CTAB) based modified RNA isolation protocol suitable for isolating high quality total RNA from different parts of field-grown jute plants was reported. Modifications come with two extra wash with extraction buffer, residual polysaccharides precipitation with absolute ethanol and potassium acetate during phenol:chloroform:isoamyl alcohol (PCI), intermediate pelleting with lithium chloride followed by dissolving the pellet in sodium dodecyl sulfate (SDS). The 28S/18S ratio and RIN of isolated RNA were greater than 2.0 and 7.3 to 8.8, respectively reveal RNA to be high purity. The isolated RNA can be used directly for subsequent downstream applications including cDNA library construction, reverse transcription polymerase chain reaction (RT-PCR) and RNA-Seq raw data generation without further purification. Moreover, this study showed that this method could also be successfully applied to other polyphenols and polysaccharides rich malvaceous plants.

 

Key words: RNA isolation, Corchorus, Jute, quantitative polymerase chain reaction with reverse transcription (RT-qPCR), cDNA library construction, next-generation sequencing (NGS) data generation.