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Evaluation of suitable reference genes for gene expression studies in the developing mouse cortex using RT-qPCR.

Ananya Uppalapati¹, Timothy Wang¹, Lena H Nguyen²

¹Department of Neuroscience, School of Behavioral and Brain Sciences, The University of Texas at Dallas, Richardson, TX, 75080, USA.

BMC Neuroscience

|February 18, 2025

Summary

For accurate gene expression analysis in developing mouse cortex, B2m, Gapdh, and Hprt are recommended reference genes for real-time quantitative PCR (RT-qPCR). These genes show stable expression during late corticogenesis, crucial for reliable neuroscience research.

Keywords:

Cerebral cortex Gene expression Housekeeping genes Mouse Neurodevelopment RT-qPCR Reference genes

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Area of Science:

Neuroscience
Molecular Biology
Genetics

Background:

Real-time quantitative PCR (RT-qPCR) is vital for neuroscience gene expression studies.
Stable reference genes are essential for accurate RT-qPCR relative quantification.
Limited data exists on reference gene stability in the developing mouse cortex.

Purpose of the Study:

To evaluate the expression stability of commonly used reference genes in the developing mouse cortex.
To identify optimal reference genes for RT-qPCR during late corticogenesis (embryonic day 15 to postnatal day 0).

Main Methods:

Assessed stability of five housekeeping genes: Actb, Gapdh, B2m, Rpl13a, and Hprt.
Utilized five computational algorithms: BestKeeper, geNorm, NormFinder, DeltaCt, and RefFinder.
Analyzed cortical tissue from mice at embryonic day 15 to postnatal day 0.

Main Results:

B2m, Gapdh, and Hprt demonstrated the most stable expression.
Combinations of B2m/Gapdh and B2m/Hprt also showed high stability.
Actb and Rpl13a exhibited the least stable expression patterns.

Conclusions:

B2m, Gapdh, and Hprt are identified as suitable reference genes for RT-qPCR in mouse cortical development studies.
These genes provide reliable normalization for gene expression analysis from embryonic day 15 to postnatal day 0.
The findings support accurate relative quantification in neuroscience research during critical developmental periods.