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Atsushi Shimizu

Atsushi Shimizu

Iwate Medical University, Japan

Title: Reduction of systematic bias in transcriptome data from human peripheral blood mononuclear cells for transportation and biobanking

Biography

Atsushi Shimizu graduated from Aoyama Gakuin University, Tokyo, Japan and obtained PhD degree in science from Aoyama Gakuin University in 1999. He then worked as a post-doc with Professor Nobuyoshi Shimizu at Keio University School of Medicine, Tokyo, Japan. He was an Associate Professor at the Keio University School of Medicine. He is now a Principal Investigator at Iwate Medical University. Over the last two years, he has developed a research project with his group focusing on the large-scale genome cohort study aiming to characterize the genome, epigenome and transcriptome analysis.

Abstract

Transportation of samples is essential for large-scale biobank projects. However, RNA degradation during pre-analytical operations prior to transportation can cause systematic bias in transcriptome data, which may prevent subsequent biomarker identification. In this study, we examined the effectiveness of RNA-stabilizing reagents to prevent RNA degradation during pre-analytical operations with an emphasis on RNA from PBMCs to establish a protocol for reducing systematic bias. To this end, we obtained PBMCs from 11 healthy volunteers and analyzed the purity, yield, and integrity of extracted RNA after performing pre-analytical operations for freezing PBMCs at -80°C. We selected 7 samples from 11 healthy volunteers, and systematic bias in expression levels was examined by RNA-Seq experiments and data analysis. Our data demonstrated that omission of stabilizing reagents significantly lowered RNA integrity, suggesting substantial degradation of RNA molecules due to pre-analytical freezing. RNA-Seq for 25,223 transcripts suggested that about 40% of transcripts were systematically biased. These results indicated that appropriate reduction in systematic bias is essential in protocols for collection of RNA from PBMCs for large-scale biobank projects. Among the seven commercially available stabilizing reagents examined in this study, RNA-Seq experiments consistently suggested that RNALock, RNA/DNA Stabilization Reagent for Blood and Bone Marrow, and 1-Thioglycerol/Homogenization solution could reduce systematic bias. On the basis of the results of this study, we established a protocol to reduce systematic bias in the expression levels of RNA transcripts isolated from PBMCs. We believe that these data provide a novel methodology for collection of high-quality RNA from PBMCs for biobank researchers