title: Recovery of cell-free mRNA and microRNA from human semen based on their physical nature.
abstract: Cell-free seminal mRNA (cfs-mRNA) and microRNA (cfs-miRNA) have been found in human ejaculate and reported as promising noninvasive biomarkers for disorders of male reproductive organs and forensic identification. However, seminal plasma is particularly challenging for RNA extraction due to its complicated composition and high content of protein, DNA, and polysaccharide. Here, we report a novel, simple, and reliable method for the isolation of cfs-mRNA and cfs-miRNA from human semen based on our previous findings of their physical nature. Seminal microvesicles (0.1-0.5 µm in diameter), which contain the majority of cfs-mRNA, were enriched by a microfilter. Protein complexes, which most cfs-miRNA is bound with, were enriched by an ultrafilter. Harvesting the complexes or microvesicles, in which RNAs exist, avoided the influence of other components in human semen, thus favoring RNA isolation and purification. This new method can efficiently isolate cfs-mRNA and cfs-miRNA separately based on their physical nature, with high RNA purity, and low DNA contamination. It may also be applied or modified to isolate cell-free RNAs in other fluids.
authors.affiliation: Family Planning Research Institute/Center of Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Liping
authors.initials: L
authors.lastname: Wang
authors.forename: Jinming
authors.initials: J
authors.lastname: Lv
authors.forename: Cuicui
authors.initials: C
authors.lastname: Guo
authors.forename: Honggang
authors.initials: H
authors.lastname: Li
authors.forename: Chengliang
authors.initials: C
authors.lastname: Xiong
mesh_terms: D005374:Filtration / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012333:RNA, Messenger / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D012661:Semen / Q000737:chemistry* / Q000166:cytology
chemical_list: D035683:MicroRNAs; D012333:RNA, Messenger
keywords: RNA purity; RNA recovery; filtration; gene expression; human ejaculate; purification