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Open Access Highly Accessed Research article

Analysis of transcript and protein overlap in a human osteosarcoma cell line

Daniel Klevebring13, Linn Fagerberg2, Emma Lundberg2, Olof Emanuelsson1, Mathias Uhlén2 and Joakim Lundeberg1*

Author affiliations

1 Science for Life Laboratory, School of Biotechnology, Division of Gene Technology, Royal Institute of Technology, SE-171 65 Solna, Sweden

2 Science for Life Laboratory, School of Biotechnology, Division of Proteomics, Royal Institute of Technology, SE-171 65 Solna, Sweden

3 Department of Medical Epidemiology and Biostatistics, Karolinska Institute, Stockholm, Sweden

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Citation and License

BMC Genomics 2010, 11:684  doi:10.1186/1471-2164-11-684

Published: 2 December 2010

Abstract

Background

An interesting field of research in genomics and proteomics is to compare the overlap between the transcriptome and the proteome. Recently, the tools to analyse gene and protein expression on a whole-genome scale have been improved, including the availability of the new generation sequencing instruments and high-throughput antibody-based methods to analyze the presence and localization of proteins. In this study, we used massive transcriptome sequencing (RNA-seq) to investigate the transcriptome of a human osteosarcoma cell line and compared the expression levels with in situ protein data obtained in-situ from antibody-based immunohistochemistry (IHC) and immunofluorescence microscopy (IF).

Results

A large-scale analysis based on 2749 genes was performed, corresponding to approximately 13% of the protein coding genes in the human genome. We found the presence of both RNA and proteins to a large fraction of the analyzed genes with 60% of the analyzed human genes detected by all three methods. Only 34 genes (1.2%) were not detected on the transcriptional or protein level with any method. Our data suggest that the majority of the human genes are expressed at detectable transcript or protein levels in this cell line. Since the reliability of antibodies depends on possible cross-reactivity, we compared the RNA and protein data using antibodies with different reliability scores based on various criteria, including Western blot analysis. Gene products detected in all three platforms generally have good antibody validation scores, while those detected only by antibodies, but not by RNA sequencing, generally consist of more low-scoring antibodies.

Conclusion

This suggests that some antibodies are staining the cells in an unspecific manner, and that assessment of transcript presence by RNA-seq can provide guidance for validation of the corresponding antibodies.