A simple and efficient method for isolation of RNA from high polyphenolic tissues of Black Pepper (Piper nigrum L.)

Abstract

Black pepper (Piper nigrum L.), member of the family Piperaceae is indigenous to India and is one of the most widely used spices in the world. Extraction of high quality, high molecular weight RNA is essential for gene expression studies in piper species. Confirmation of apomixis and identification of apomictic related genes could be of great importance in pepper breeding. For the detection of apomictic gene expression, isolation of RNA from berries is essential. The berries of black pepper contain high levels of storage lipids, storage proteins, polysaccharides, and polyphenolics which often hinder extraction of high quality RNA. The studies on functional genomic aspects in black pepper is limited due to high polyphenol contamination in routine RNA extraction procedures, which further hampers crop improvement programme. In the present study to analyse the transcriptome, extraction of high quality RNA from mature berries was carried out. The commercially available RNA isolation kits that use guanidinium salt solutions were ineffective in yielding significant amounts of high quality RNA. Here, we report for the first time a simple and efficient method for isolating high quality RNA from berries of piper nigrum using soluble polyvinylpyrrolidone (PVP) and precipitation of RNA by lithium chloride, which was a successful approach in extracting high quality RNA for further gene expression analysis by DDRT PCR, SSH etc.