A study on total RNA including microRNA isolation from cell lines by different cost effective methods
Abstract
Small non-coding RNAs have been recognized as potential controllers of various biological functions, including gene expression, molecular growth, and physiological functions. The diverse ways considerably aid the prognosis and diagnosis of many diseases which microRNAs (miRNAs) manifest themselves. It is crucial to successfully isolate miRNAs from various tissues and bodily fluids since they have a wide range of uses as clinical biomarkers for the precise detection of many diseases, cancer risk assessment, and investigation of different metabolic processes and gene activity. The present study compares the available RNA extraction protocols and renders an improvised protocol for isolating RNA and subsequent extraction of miRNA molecules. Different RNA isolation protocols were analyzed: Direct-zol™ RNA MiniPrep Plus kit, TRIzolTM reagent, and non-kit organic RNA extraction methods. The quality of RNA was assessed using spectrophotometry and bleach gel electrophoresis. Here, we have measured the expression of miRNA10b, a specific biomarker candidate for the prognosis of colorectal cancer that plays a major role in metastasis. The amplified expression of miRNA was demonstrated using Real-time PCR. The qualitative and quantitative analysis of the extracted RNA was evaluated. The modified extraction protocol was found to yield good-qualitymiRN. The modified RNA extraction protocol is a simple, cost-effective, efficient method for miRNA isolation from cell lines.
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