Abstract
For many years real-time quantitative polymerase chain reaction (qPCR) has been the golden standard to measure gene expression levels in brain tissue. However, today it is generally accepted that many factors may affect the outcome of the study and more consensus is required to perform and interpret real-time qPCR experiments in a comparable way. Here we describe the basic techniques used for more than a decade in our laboratory to extract RNA and protein from the same piece of frozen brain tissue and to quantify relative mRNA levels with real-time qPCR and SYBR Green.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Elfving B, Bonefeld BE, Rosenberg R et al (2008) Differential expression of synaptic vesicle proteins after repeated electroconvulsive seizures in rat frontal cortex and hippocampus. Synapse 62(9):662–670. https://doi.org/10.1002/syn.20538
Bustin SA, Benes V, Garson JA et al (2009) The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55(4):611–622. https://doi.org/10.1373/clinchem.2008.112797
Bustin S, Nolan T (2017) Talking the talk, but not walking the walk: RT-qPCR as a paradigm for the lack of reproducibility in molecular research. Eur J Clin Investig 47(10):756–774. https://doi.org/10.1111/eci.12801
Bonefeld BE, Elfving B, Wegener G (2008) Reference genes for normalization: a study of rat brain tissue. Synapse 62(4):302–309. https://doi.org/10.1002/syn.20496
Elfving B, Müller HK, Oliveras I et al (2019) Differential expression of synaptic markers regulated during neurodevelopment in a rat model of schizophrenia-like behavior. Prog Neuro-Psychopharmacol Biol Psychiatry 95:109669. https://doi.org/10.1016/j.pnpbp.2019.109669
Tian Q, Stepaniants SB, Mao M et al (2004) Integrated genomic and proteomic analyses of gene expression in mammalian cells. Mol Cell Proteomics 3(10):960–969. https://doi.org/10.1074/mcp.M400055-MCP200
Pradet-Balade B, Boulme F, Beug H et al (2001) Translation control: bridging the gap between genomics and proteomics? Trends Biochem Sci 26(4):225–229. https://doi.org/10.1016/s0968-0004(00)01776-x
Müller HK, Wegener G, Popoli M et al (2011) Differential expression of synaptic proteins after chronic restraint stress in rat prefrontal cortex and hippocampus. Brain Res 1385:26–37. https://doi.org/10.1016/j.brainres.2011.02.048
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Elfving, B. (2022). Investigation of Synaptic Vesicle Proteins in Rat Brain Tissue Using Real-Time qPCR. In: Dahlmanns, J., Dahlmanns, M. (eds) Synaptic Vesicles. Methods in Molecular Biology, vol 2417. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1916-2_5
Download citation
DOI: https://doi.org/10.1007/978-1-0716-1916-2_5
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1915-5
Online ISBN: 978-1-0716-1916-2
eBook Packages: Springer Protocols