This webinar will address improvements in the library prep workflow for small RNA sequencing in serum and plasma. Sequencing of small RNAs from extracellular fluids, including serum and plasma, is a fast-growing area of biomarker research. Analysis of plasma and serum by small RNA sequencing involves isolating RNA, performing library preparation, sequencing, and downstream data analysis.
Our speaker, David Simpson of Queens University Belfast, will describe the factors to be considered at each of these steps and will compare workflows from Qiagen and Bioo Scientific/PerkinElmer for small RNA sequencing from serum and plasma samples.
The workflow from Bioo Scientific involves magnetic bead-based RNA purification followed by library preparation using adapters with randomized ends to reduce ligation bias, while the Qiagen workflow involves column-based purification of RNA followed by library preparation utilizing unique molecular identifiers for PCR bias correction. Dr. Simpson will discuss metrics such as mapping rate, adapter-dimer contamination, and diversity of miRNAs detected.
Dr. Simpson will also discuss the effectiveness of the alternative chemistries and randomized adapters employed by Bioo Scientific to reduce ligation bias. For example, the correction of PCR bias by use of unique molecular identifiers in the Qiagen workflow had little effect upon relative miRNA quantification, suggesting that reduction of ligation bias is a more effective strategy to reduce bias in small RNA library preparation.
Adam R. Morris obtained his PhD in Genetics and Genomics from Duke University in 2010 in the lab of Dr. Jack Keene. He then did postdocs at the Netherlands Cancer Institute (NKI) with Dr. Reuven Agami and at the University of Texas at Austin with Dr. Vishy Iyer before joining Bioo Scientific, a PerkinElmer company, in 2013.
Queens University Belfast
Dr Simpson obtained a BSc (Hons) in Biological Sciences (1988) and PhD in Molecular microbiology (1995) at the University of Leicester. He held several postdoctoral positions at Queen’s University Belfast, where he is now a Senior Lecturer in the Centre for Experimental Medicine. His group were amongst the first to exploit next generation sequencing (NGS) techniques to screen panels of candidate genes for the inherited retinal disease Retinitis Pigmentosa (RP). Another area of interest is the molecular cell biology underlying diabetic vascular complications, specifically the role of microRNAs in regulating gene expression in the retina and vasculature. He is currently using NGS to profile small RNAs in model systems and in blood as potential prognostic markers for cardiovascular disease.