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Published on: February 13, 2013
Selection and Validation of Reference Genes for Pan-Cancer in Platelets Based on RNA-Sequence Data
Xiaoxia Wen1, Guishu Yang2, Yongcheng Dong3
1Chongqing Key Laboratory of Sichuan-Chongqing Co-construction for Diagnosis and Treatment of Infectious Diseases Integrated Traditional Chinese and Western Medicine, College of Medical Technology, Chengdu University of Traditional Chinese Medicine, Chengdu, China.
This study identified Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as the most stable reference gene in platelets for pan-cancer diagnosis. Accurate mRNA analysis in platelets is crucial for early cancer detection.
Area of Science:
- Molecular Biology
- Oncology
- Biomarker Discovery
Background:
- Platelet messenger RNA (mRNA) shows potential as a pan-cancer biomarker.
- Quantitative real-time polymerase chain reaction (RT-qPCR) is a key technique for measuring mRNA expression.
- Reliable reference genes are essential for accurate RT-qPCR analysis, but none are validated for pan-cancer in platelets.
Purpose of the Study:
- To identify and validate the most suitable reference gene for platelet mRNA expression analysis in pan-cancer.
- To ensure the accuracy and reliability of RT-qPCR-based diagnostic platforms for early cancer detection.
Main Methods:
- Bioinformatic analysis of platelet RNA-sequencing data (GSE68086) to identify 95 candidate reference genes.
- Screening and verification of seven candidate genes (YWHAZ, GNAS, GAPDH, OAZ1, PTMA, B2M, ACTB) using another dataset (GSE89843).
- RT-qPCR validation of candidate genes in pan-cancer patients and healthy individuals, followed by stability analysis using comparative Delta CT, geNorm, NormFinder, and BestKeeper.
Main Results:
- Bioinformatic analysis yielded 95 candidate reference genes, from which seven were selected for further validation.
- RT-qPCR and stability analysis revealed that GAPDH exhibited the most stable and constitutive high-level expression across all tested samples.
- Other commonly used reference genes showed variable expression, highlighting the need for specific validation.
Conclusions:
- Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is recommended as the optimal reference gene for platelet transcript analysis in pan-cancer studies.
- The use of validated reference genes like GAPDH will enhance the reliability of platelet-based molecular diagnostic platforms for early cancer detection.
