RNA-seq reveals more consistent reference genes for gene expression studies in human non-melanoma skin cancers

Van L T Hoang; Lisa N Tom; Xiu-Cheng Quek; Jean-Marie Tan; Elizabeth J Payne; Lynlee L Lin; Sudipta Sinnya; Anthony P Raphael; Duncan Lambie; Ian H Frazer; Marcel E Dinger; H Peter Soyer; Tarl W Prow

doi:10.7717/peerj.3631

RNA-seq reveals more consistent reference genes for gene expression studies in human non-melanoma skin cancers

PeerJ. 2017 Aug 21:5:e3631. doi: 10.7717/peerj.3631. eCollection 2017.

Authors

Van L T Hoang^#¹, Lisa N Tom^#¹, Xiu-Cheng Quek^#^{2

3}, Jean-Marie Tan¹, Elizabeth J Payne¹, Lynlee L Lin¹, Sudipta Sinnya¹, Anthony P Raphael^{1

4}, Duncan Lambie⁵, Ian H Frazer⁶, Marcel E Dinger^{2

3}, H Peter Soyer¹, Tarl W Prow^{1

7}

Affiliations

¹ Dermatology Research Centre, Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, The University of Queensland, Brisbane, Queensland, Australia.
² Garvan Institute of Medical Research, Sydney, New South Wales, Australia.
³ St Vincent's Clinical School, University of New South Wales, Sydney, New South Wales, Australia.
⁴ Wellman Center for Photomedicine, Harvard Medical School, Massachusetts General Hospital, Boston, MA, USA.
⁵ Department of Anatomical Pathology, Princess Alexandra Hospital, Brisbane, Queensland, Australia.
⁶ Diamantina Institute, Translational Research Institute, Princess Alexandra Hospital, The University of Queensland, Brisbane, Queensland, Australia.
⁷ Biomaterials Engineering and Nanomedicine Strand, Future Industries Institute, University of South Australia, Mawson Lakes, Australia.

^# Contributed equally.

Abstract

Identification of appropriate reference genes (RGs) is critical to accurate data interpretation in quantitative real-time PCR (qPCR) experiments. In this study, we have utilised next generation RNA sequencing (RNA-seq) to analyse the transcriptome of a panel of non-melanoma skin cancer lesions, identifying genes that are consistently expressed across all samples. Genes encoding ribosomal proteins were amongst the most stable in this dataset. Validation of this RNA-seq data was examined using qPCR to confirm the suitability of a set of highly stable genes for use as qPCR RGs. These genes will provide a valuable resource for the normalisation of qPCR data for the analysis of non-melanoma skin cancer.

Keywords: Non-melanoma skin cancer; RNA-seq; Reference gene; qPCR.

Grants and funding

This project was supported by Epiderm (Boronia Park, NSW 2111, Australia) and NHMRC Fellowships APP1109749 and APP1088318. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.