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RPS13, a potential universal reference gene for normalisation of gene expression in multiple human normal and cancer tissue samples

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Abstract

Background

Reference genes are considered stable genes and are used for normalizing the gene expression profile across different cell types; as well as, in normal and diseased samples. However, these gene associates with different biological processes, and hence expression vary in different pathological conditions. Therefore, in the present study, eight different reference genes were used and compared to identify common reference gene usable for an array of different cell types and human cancers.

Methods and results

The expression stability of the eight reference genes across eleven normal and cancerous tissues was confirmed through real time-qPCR. Ribosomal protein S13 (RPS13) was found to be a common and stable reference gene across intra- and inter-comparison between various normal and tumor tissue types. Further, TCGA data analysis across and between normal and tumor tissue types also showed minimum deviation in expression of RPS13 gene out of eight routinely used reference genes.

Conclusion

RPS13 is the common stable reference gene in normalization for gene expression based analysis in cancer research.

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Data availability

The raw data and/or analyzed data are available from the corresponding and first authors on reasonable request.

Code availability

Not applicable.

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Acknowledgements

Authors are grateful to all members of Gupta Lab, ACTREC for valuable discussions. This work was partly supported by TMC-Intramural Research Grant. We are thankful to Dr. Manisha Kulkarni and Mr. Anand Deshpande from TMH for their kind co-operation in the collection of tissue samples from TMH-TTR.

Funding

This research was partly supported by TMC-Intramural Research Grant.

Author information

Author notes

  1. Mudasir Rashid and Sanket Girish Shah are regarded as joint first authors and have contributed equally to this work.

Authors and Affiliations

  1. Epigenetics and Chromatin Biology Group, Gupta Lab, Cancer Research Institute, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Kharghar, Navi Mumbai, MH, 410210, India

    Mudasir Rashid, Sanket Girish Shah, Abhiram Natu, Tripti Verma, Sukanya Rauniyar & Sanjay Gupta

  2. Homi Bhabha National Institute, Training School Complex, Anushakti Nagar, Mumbai, MH, India

    Mudasir Rashid, Sanket Girish Shah, Abhiram Natu, Tripti Verma, Sukanya Rauniyar & Sanjay Gupta

  3. Biorepository, Advanced Centre for Treatment, Research and Education in Cancer, Tata Memorial Centre, Kharghar, Navi Mumbai, MH, 410210, India

    Poonam B. Gera

Authors
  1. Mudasir Rashid
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Contributions

SG conceived the idea, designed the experiments, and wrote the manuscript. MR performed the majority of experiments with contributions from SGS, AN, TV, and SR. All authors contributed to manuscript writing. SGS contributed to the designing of experiments and major writing of the manuscript. AN performed data analysis. PG validated H&E stained slides. SG and SGS revised the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Sanjay Gupta.

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Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

The project was approved by the Institute human ethics committee vide #164 dated 27-04-2015.

Consent for publication

Waiver of consent was granted for retrospective study proposal by Institutional Ethics Committee.

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Supplementary Information

Below is the link to the electronic supplementary material.

11033_2021_6828_MOESM1_ESM.tif

Supplementary file1 Fig. S1 Histopathological analysis of human tissue samples after haematoxylin and eosin staining. Representative images for (a-d) Adenocarcinoma (stomach, colon, rectum and ovary) cancer type; (e-g) Squamous cell carcinoma (tongue, buccal and penis) cancer type; (h-k) Others represent glioblastoma, hepatocellular carcinoma, renal cell carcinoma and invasive ductal carcinoma (TIF 9573 kb)

11033_2021_6828_MOESM2_ESM.tif

Supplementary file2 Fig. S2 Stability prediction of reference genes across normal and tumor tissue types (A) NormFinder stability prediction of housekeeping genes across normal tissue types (B) NormFinder stability prediction of housekeeping genes across tumor tissue types (TIF 4416 kb)

11033_2021_6828_MOESM3_ESM.tif

Supplementary file3 Fig. S3 Comprehensive stability prediction of reference genes via Delta CT, BestKeeper, Normfinder and Genorm (A) Ranking order of housekeeping genes across tumor tissue types (B) Stability value of reference genes (TIF 2967 kb)

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Rashid, M., Shah, S.G., Natu, A. et al. RPS13, a potential universal reference gene for normalisation of gene expression in multiple human normal and cancer tissue samples. Mol Biol Rep 48, 7967–7974 (2021). https://doi.org/10.1007/s11033-021-06828-6

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  • DOI: https://doi.org/10.1007/s11033-021-06828-6

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