02097nas a2200253   4500000000100000000000100001008004100002260001500043653003900058653003000097653002000127653000800147653001900155100001500174700002100189700002300210700002300233245008600256856004600342300001000388490000600398520142500404022001401829       2016                            d  c2016-08-0910aExtracellular signalling molecules10aHigh-throughput screening10aNon-coding RNAs10aRNA10aRNA sequencing1 aZhiyun Wei1 aArsen O. Batagov1 aDavid R. F. Carter1 aAnna M. Krichevsky00aFetal Bovine Serum RNA Interferes with the Cell Culture derived Extracellular RNA  uhttps://www.nature.com/articles/srep31175  a311750 v63 aFetal bovine serum (FBS) has been used in eukaryotic cell cultures for decades. However, little attention has been paid to the biological effects associated with RNA content of FBS on cell cultures. Here, using RNA sequencing, we demonstrate that FBS contains a diverse repertoire of protein-coding and regulatory RNA species, including mRNA, miRNA, rRNA and snoRNA. The majority of them (>70%) are retained even after extended ultracentrifugation in the preparations of vesicle-depleted FBS (vdFBS) commonly utilized in the studies of extracellular vesicles (EV) and intercellular communication. FBS-associated RNA is co-isolated with cell-culture derived extracellular RNA (exRNA) and interferes with the downstream RNA analysis. Many evolutionally conserved FBS-derived RNA species can be falsely annotated as human or mouse transcripts. Notably, specific miRNAs abundant in FBS, such as miR-122, miR-451a and miR-1246, have been previously reported as enriched in cell-culture derived EVs, possibly due to the confounding effect of the FBS. Analysis of publically available exRNA datasets supports the notion of FBS contamination. Furthermore, FBS transcripts can be taken up by cultured cells and affect the results of highly sensitive gene expression profiling technologies. Therefore, precautions for experimental design are warranted to minimize the interference and misinterpretations caused by FBS-derived RNA.  a2045-2322