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Open Access Highly Accessed Research article

Increased sensitivity of next generation sequencing-based expression profiling after globin reduction in human blood RNA

Anastasios Mastrokolias1, Johan T den Dunnen12, GertJan B van Ommen1, Peter AC 't Hoen1 and Willeke MC van Roon-Mom1*

Author Affiliations

1 Center for Human and Clinical Genetics, Leiden University Medical Center, Einthovenweg 20, 2333ZC, Leiden, The Netherlands

2 Leiden Genome Technology Center, Leiden University Medical Center, Einthovenweg 20, 2333ZC Leiden, The Netherlands

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BMC Genomics 2012, 13:28  doi:10.1186/1471-2164-13-28

Published: 18 January 2012

Abstract

Background

Transcriptome analysis is of great interest in clinical research, where significant differences between individuals can be translated into biomarkers of disease. Although next generation sequencing provides robust, comparable and highly informative expression profiling data, with several million of tags per blood sample, reticulocyte globin transcripts can constitute up to 76% of total mRNA compromising the detection of low abundant transcripts. We have removed globin transcripts from 6 human whole blood RNA samples with a human globin reduction kit and compared them with the same non-reduced samples using deep Serial Analysis of Gene Expression.

Results

Globin tags comprised 52-76% of total tags in our samples. Out of 21,633 genes only 87 genes were detected at significantly lower levels in the globin reduced samples. In contrast, 11,338 genes were detected at significantly higher levels in the globin reduced samples. Removing globin transcripts allowed us to also identify 2112 genes that could not be detected in the non-globin reduced samples, with roles in cell surface receptor signal transduction, G-protein coupled receptor protein signalling pathways and neurological processes.

Conclusions

The reduction of globin transcripts in whole blood samples constitutes a reproducible and reliable method that can enrich data obtained from next generation sequencing-based expression profiling.