Improve sample preparation process for miRNA isolation from the culture cells by using silica fiber membrane

Wen Pin Hu, Yu Chi Chen, Wen Yih Chen

研究成果: 雜誌貢獻期刊論文同行評審

3 引文 斯高帕斯(Scopus)

摘要

In clinical applications of miRNAs, the purity and quality of the testing samples are very critical, especially the obtained tissue sample volume is limited. If the extracted miRNAs are contaminated or different in quality before analysis, it will increase the variance of the analysis result and make the medical information judgment incorrect and cannot be portable. Herein, we improved the commercially extraction kit by realizing the fundamental mechanism and hoped to serve finding optimal procedures for increasing the recovery of miRNAs extracted from cultured cells. In the adsorption process, the factors, like increasing the ethanol concentration or adding Ca2+, could influence the RNA adsorption were investigated. For the elution process, the effect caused by raising the elution temperature and raising the pH value of elution buffer was studied. Finally, the conditions for miRNA extraction are optimal modified by using a 65% (v/v) solution of ethanol in the adsorption process, and using TE buffer with the pH value of 8.0 and raising the temperature to 55 °C in the elution. According to the quantified results, the improved extraction kit can promote the recovery of endogenous miR-21 by about 6 times by using the optimal extraction conditions comparing with the miRNeasy Mini Kit.

原文???core.languages.en_GB???
文章編號21132
期刊Scientific Reports
10
發行號1
DOIs
出版狀態已出版 - 12月 2020

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