ABSTRACTCurrent RNA purification methods widely use silica-based columns that allow quick isolation of high quality and right quantities of RNA. However, the major limitations include high cost, the requirement of different kits for small RNA isolation, genomic DNA contamination, and not being flexible. Here, we used the in-house RNA isolation reagent for cell lysis, followed by precipitation of RNA using isopropanol resulted in a similar quantity and quality of RNA compared to the commercial TRIzol. The commercial RNA isolation kits with silica-based columns recommend genomic DNA digestion during or after RNA purification adding time and cost to RNA purification. Here, we developed an optimized in-house protocol for isolating high-quality RNA free of genomic DNA contamination using magnetic silica beads without the need for DNase digestion. Additionally, our method purifies total RNA along with the small RNA fraction, including miRNAs, which usually require a separate kit for extraction. Additionally, the RNA prepared with our method was equally suitable for mRNA and miRNA expression analysis using RT-qPCR. Together, the in-house method of RNA isolation using a magnetic bead exhibited comparable or better total RNA extraction compared to commercial kits at a fraction of the cost and across various cells and tissues.
Small RNA Isolation from Tissues of Grapevine and Woody Plants