胎牛血清RNA干扰了细胞培养外源性RNA

几年前提出，并在该领域仍然高度争议的一个重要问题是，是否特异性RNA种类或图案在电动汽车和的RNP充实。这种富集将支持RNA分泌的积极和监管机制?？商娲?，exRNA可以反映整个蜂窝转录，特定的RNA类（如miRNA）的，或RNA的代谢的步骤之一，因此，对于监视蜂窝转录提供一个外围设备的措施。解决这个问题是细胞间通讯和RNA的生物标志物的发展研究是至关重要的。细胞培养系统exRNA生物合成的研究使用和释放，在很大程度上依赖于胎牛血清（FBS），常用的试剂众多类型细胞的生长至关重要。尽管许多报道特点exRNA从人类和小鼠血清中分离和血清建议RNA作为疾病生物标志的主要来源1，很少有人注意支付给FBS衍生的RNA的基于细胞培养研究混杂效应。在这里，我们描述的FBS的RNA含量以及囊泡-贫化的FBS（vdFBS），并证明其与细胞衍生exRNA从条件培养基，可能的话，细胞RNA分离的干扰。

英文原文：

Fetal  Bovine Serum RNA Interferes with the Cell Culture derived Extracellular RNA

Abstract

Fetal 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.

Introduction

Deep sequence analysis of extracellular RNA (exRNA) released by cultured cells in forms of extracellular vesicles (EVs) and lipoprotein complexes (RNPs) is an expanding area of research. An important question raised years ago and still highly debated in the field, is whether specific RNA species or motifs are enriched in EVs and RNPs.