housekeepingGene

Housekeeping genes are genes that encode proteins required for basic cellular maintenance and are expected to be expressed at relatively constant levels across many tissues and conditions. They typically participate in essential processes such as metabolism, transcription, translation, and structural maintenance, making them useful internal controls for measuring gene expression. In practice, housekeeping genes are used to normalize data in experiments like quantitative PCR and RNA sequencing to correct for variation in RNA input, quality, or reverse transcription efficiency.

Commonly used housekeeping genes include GAPDH, ACTB (beta-actin), 18S ribosomal RNA, RPLP0, B2M, and HPRT1, among others. However, no single gene remains perfectly stable in all contexts. Expression of housekeeping genes can vary with tissue type, developmental stage, disease state, treatment, and experimental conditions, which can bias normalization if relied on without validation. Therefore, candidate reference genes should be validated for stability in the specific study.

Best practices involve using multiple reference genes and assessing their stability with dedicated tools or algorithms, such as geNorm, NormFinder, or BestKeeper. It is advisable to select genes that are not co-regulated with targets of interest and to adjust normalization strategy accordingly. In some contexts, especially high-throughput analyses, alternative normalization methods that do not depend on a single reference gene may be preferred, though housekeeping genes remain a common normalization option for targeted assays.