Method for separating low-molecular weight ribonucleic acid (RNA) of plant

Abstract

The invention provides a method for separating low-molecular weight ribonucleic acid (RNA) of plant. The method mainly comprises the following steps of: treating a sample, removing deoxyribonucleic acid (DNA) and protein, removing high-molecular weight RNA, obtaining the low-molecular weight RNA, and storing the low-molecular weight RNA. In the extraction process of the low-molecular weight RNA, the DNA, the high-molecular weight RNA and the low-molecular weight RNA are precipitated at different stages, the DNA and the high-molecular weight RNA are precipitated by using 30 percent polyethylene glycol (PEG) and absolute ethanol in an amount which is 1/3 the volume of a tube, and the low-molecular weight RNA is precipitated by using absolute ethanol in an amount which is 2/3 volume of the tube, so that the low-molecular weight RNA is enriched; and the extraction process is easy and convenient to operate, the required time is short, and the method is a simple, economic and efficient method for extracting high-quality low-molecular weight RNA of the plant.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a method for isolating plant low-molecular-weight RNA. Background technique [0002] Low-molecular-weight RNAs include siRNAs and miRNAs with sizes ranging from 20nt to 24nt. These small RNA molecules play an important regulatory role in plant growth, development and response to adversity stress. Since scientists first discovered miRNAs in plants in 2002, the miRNAs database has included 4169 plant miRNAs, and the research on small RNA molecules has attracted more and more attention from plant scientists. The acquisition of small RNA molecules is the prerequisite for carrying out small RNA research, and the quality of small RNA directly affects the development of subsequent experiments. No matter which extraction method is used, it is required that the obtained low-molecular-weight RNA has high purity and good quality, without contamination by polyphenols, ...

AI Technical Summary

Problems solved by technology

Small RNA extraction kits developed by companies such as Ambion and Stratagene use membranes made of polymer materials to absorb small RNAs, and then recover them from the membranes. Although this method is convenient to use, it is expensive, and these kits Not suitable for large-scale extraction of small RNA

The method of recovering small RNA from total RNA is a classic method of isolating small RNA. This method first isolates total RNA (large molecular weight RNA and low molecular weight RNA) from plant tissues, and then uses the method of gel cutting to recover from total RNA. Recovery of low-molecular-weight RNA in medium and long-term methods, and the yield of small RNA is low

There are two methods for directly enriching small RNAs reported in the literature, one is to use 7.5% polyethylene glycol and 1.25mol / L sodium chloride to precipitate large molecular weight RNA, and then use an equal volume of isopropanol to precipitate and recover overnight small RNA; the other is to use 5% polyethylene glycol and 1 / 9 volume of isopropanol to precipitate large molecular weight RNA, and then use an equal volume of isopropanol to precipitate small RNA. The rate is high, but the time-consuming is relatively long

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