Enhanced extraction efficiency of miRNA from cells by addition of Triton X-100

Abstract

The determination of microRNA (miRNA) levels in biomaterials has become important for understanding their biological functions and for the diagnosis of various diseases. An effective extraction method is needed for maximizing the recovery of miRNAs from cells, while minimizing RNA degradation during the extraction because miRNAs present only approximately 0.01 % of total RNA. In this study, we used Triton X-100 (TX-100) to improve the extraction efficiency of miRNAs with TRIzol® reagent, which is a commonly used commercial microRNA isolation kit. The concentration of TX-100 and the incubation time after the addition of TX-100 were optimized to maximize the extraction efficiency. The extraction recovery by a combination of TX-100 and TRIzol® reagent was approximately 1.9-fold greater than that by the TRIzol® reagent alone. We have established a very effective extraction method for the extraction of low-abundance miRNAs in biological samples for the determination of miRNA levels in biomaterials.

http://pierre.fkit.hr/hdki/cabeq/pdf/28_3_2014/Cabeq%202014-03-10.pdf

“…11 Previous studies had shown that Triton X-100 had a specific dissolving effect on the cyto- In this study, several chemical treatment techniques were verified for releasing cytoplasmic cephalosporin C deacetylase (CAH) from recombinant Escherichia coli (E. coli) cells. Among all the chemical treatment methods, using cationic detergent, organic solvents and chelating agents as permeabilizing reagents exhibited limited capacities for extraction of cytoplasmic CAH from cells.…”

Synthesis of tetracycline-imprinted polymer microspheres by reversible addition-fragmentation chain-transfer precipitation polymerization using polyethylene glycol as a coporogen

“…Reversible addition-fragmentation chain-transfer (RAFT) polymerization is a type of CRP with merits of wide application and mild reaction conditions [16]. RAFT polymerization has been used in MIP synthesis such as thin films, nanoparticles, monolithic columns, and restricted access materials [17][18][19][20]. It has been found that microspheres with uniform particle size distribution can be obtained more easily using RAFT polymerization [21].…”

ABC Spotlight on emerging microRNA analysis methods

“…A key challenge in the detection of miRNAs is sample preparation because miRNA represents a very small part of the RNA content in biological samples; for example, miRNA comprises about 0.01% of the total RNA in cells [3]. A 2-fold increase in the quantity of miRNA recovered from cells was reported when a nonionic surfactant (Triton X-100) was used in conjunction with the standard detergents in a commercial miRNA preparation system (TRIzol, Life Technologies, Grand Island, NY) [3].…”

“…A 2-fold increase in the quantity of miRNA recovered from cells was reported when a nonionic surfactant (Triton X-100) was used in conjunction with the standard detergents in a commercial miRNA preparation system (TRIzol, Life Technologies, Grand Island, NY) [3]. The improved miRNA extraction was verified through capillary electrophoresis analysis of miRNA-21 extracted from H9c2 and HeLa cells.…”