High Efficiency RNA Extraction From Sperm Cells Using Guanidinium Thiocyanate Supplemented With Tris(2-Carboxyethyl)Phosphine.

ABSTRACT: The extraction of high-quality ribonucleic acid (RNA) from tissues and cells is a key step in many biological assays. Guanidinium thiocyanate-phenol-chloroform (AGPC) is a widely used and efficient method to obtain pure RNA from most tissues and cells. However, it is not efficient with some cells like sperm cells because they are resistant to chaotropic lysis solutions containing guanidinium thiocyanate such as Buffer RLT+ and Trizol. Here, we show that disulfide bonds are responsible for the chemical resistance of sperm cells to RNA extraction reagents. We show that while β-mercaptoethanol (βME) can increase sperm lysis in Buffer RLT+, it has no effect in Trizol and leaves sperm cells intact. We measured the reduction of disulfide bonds in 2,2'-dithiodipyridine (DTDP) and observed that βM

SUBMITTER: Roszkowski M 

PROVIDER: S-EPMC8097045 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature