RNA-seq sample preparation for MiSeq sequencing (Illumina) using SPIA amplification method

 
LEAD AUTHOR

Na Li
PhD candidate
Singapore-MIT Alliance for Research and Technology
Singapore

linafromsmart@gmail.com

OTHER AUTHORS:

Bevin P. Engelward
Professor, Department of Biological Engineering
Massachusetts Institute of Technology
Cambridge, MA, USA


PROTOCOL TYPE: Real Methods
Protocol Document
SYNOPSIS:

RNA extraction from tissue or low yield samples and preparation of RNA samples for RNA-sequencing (RNA-seq) using NuGEN’s SPIA (Single Primer Isothermal Amplification) amplification technology.


ABSTRACT:

RNA-seq has become a very useful tool for transcriptome sequencing and quantification. It has also been successfully applied in the whole genome sequencing for RNA viruses. However, common methods of RNA-seq prep require huge starting quantity of high-quality RNA. In order to tackle the problem of working with biological samples with low RNA yield or compromised quality (due to degradation), we used NuGEN’s SPIA (Single Primer Isothermal Amplification) amplification method to amplify cDNA. This method increases copy numbers while avoiding the common pitfall of accumulating DNA polymerase errors in polymerase chain reaction.


 

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