Method for extracting micro ribonucleic acid from serum

Abstract

The invention provides a method for extracting micro ribonucleic acid (miRNA) from serum. The method comprises the steps that: 500[mu]l of TRIzol is added to 500[mu]l of blood plasma or serum, the mixture is violently concussed for 30s by using a vortex oscillator, and the mixture is then stood under room temperature for 5min; 100[mu]l of isopropanol is added to the mixture, the mixture is well-mixed by tumbling, then the mixture is violently concussed for 30s by using a vortex oscillator until the liquid turns transparent, and the liquid is stood under room temperature for 5min; the liquid is separated for 10min by using a centrifuge under a temperature of 4 DEG C with a speed of 3000rpm, and the supernatant is transferred into a new centrifuge tube; 500[mu]l of chloroform is added to the supernatant in the centrifuge tube, and the liquid is well-mixed by tumbling, then the liquid is separated for 10min by using a centrifuge under a temperature of 4 DEG C with a speed of 13000rpm, and the supernatant is transferred into a new centrifuge tube; ethanol with a volume which is 2.5 times than the supernatant in the centrifuge tube is added to the centrifuge tube, the liquid is well-mixed by tumbling, and the liquid is then stood under room temperature for 10min; the liquid is separated for 10min by using a centrifuge under a temperature of 4 DEG C with a speed of 13000rpm, and then RNA is precipitated; ethanol is removed by absorption, and RNA is dried; the obtained RNA is dissolved with 30[mu]l of DEPC-treated water, and the solution is processed through water-bath for 10min at a temperature of 58 DEG C, such that a target extract is obtained. The method provided by the present invention is simple and applicable. With the method, serum miRNA loss during extraction can be avoided. The method also has advantages of low extraction cost, fast speed and high extraction rate.

Description

technical field [0001] The invention belongs to a method for extracting microRNA from serum. Background technique [0002] miRNA (microRNA micro ribonucleic acid) is a type of endogenous single-stranded non-coding RNA (ribonucleic acid), generally composed of 21 nucleotides. The Sanger miRNA database version 16.0 released on September 10, 2010 contains 17,341 mature miRNA sequences from 142 species, including 1,048 miRNAs that have been cloned from human cells (http: / / www.mirbase.org) . As micro-regulatory molecules, miRNA specifically binds to the 3'-UTR of mRNA (messenger ribonucleic acid), inhibits protein translation or directly causes mRNA degradation, and regulates protein expression at the post-transcriptional level. Bioinformatics prediction results show that the translation activity of about 50% of protein-coding genes in mammals is regulated by miRNA. Functional studies have shown that miRNAs are involved in the regulation of almost all cell life processes, and ...

AI Technical Summary

Problems solved by technology

For example, miRNA is much smaller than mRNA in length, and miRNA mainly exists in the form of protein complexes in cells; conventional TRIzol extraction methods will cause a large loss of serum miRNA

The separation column extraction method for miRNA can solve the problem to a certain extent, but the adsorption of the column material will also cause the loss of miRNA, and the cost of the separation column method is high, and the price of extracting a sample is generally about 10 times that of the TRIzol method

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