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Integrated RNA extraction and RT-PCR for semi-quantitative gene expression studies on a microfluidic device

Shaw, Kristy J., Hughes, Elizabeth M., Dyer, Charlotte E., Greenman, John and Haswell, Stephen J. 2013, Integrated RNA extraction and RT-PCR for semi-quantitative gene expression studies on a microfluidic device, Laboratory investigation, vol. 93, pp. 961-966, doi: 10.1038/labinvest.2013.76.

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Title Integrated RNA extraction and RT-PCR for semi-quantitative gene expression studies on a microfluidic device
Author(s) Shaw, Kristy J.
Hughes, Elizabeth M.
Dyer, Charlotte E.
Greenman, John
Haswell, Stephen J.
Journal name Laboratory investigation
Volume number 93
Start page 961
End page 966
Total pages 6
Publisher United States and Canadian Academy of Pathology
Place of publication Baltimore, Md.
Publication date 2013
ISSN 0023-6837
Keyword(s) diagnostic
Summary This paper describes the development of a microfluidic methodology, using RNA extraction and reverse transcription PCR, for investigating expression levels of cytochrome P450 genes. Cytochrome P450 enzymes are involved in the metabolism of xenobiotics, including many commonly prescribed drugs, therefore information on their expression is useful in both pharmaceutical and clinical settings. RNA extraction, from rat liver tissue or primary rat hepatocytes, was performed using a silica-based solid-phase extraction technique. Following elution of the purified RNA, amplification of target sequences for the housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the cytochrome P450 gene CYP1A2, was carried out using a one-step reverse transcription PCR. Once the microfluidic methodology had been optimized, analysis of control and 3-methylcholanthrene-induced primary rat hepatocytes were used to evaluate the system. As expected, GAPDH was consistently expressed, whereas CYP1A2 levels were found to be raised in the drug-treated samples. The proposed system offers an initial platform for development of both rapid throughput analyzers for pharmaceutical drug screening and point-of-care diagnostic tests to aid provision of drug regimens, which can be tailor-made to the individual patient.
Language eng
DOI 10.1038/labinvest.2013.76
Field of Research 060101 Analytical Biochemistry
Socio Economic Objective 920203 Diagnostic Methods
HERDC Research category C1.1 Refereed article in a scholarly journal
Copyright notice ©2013, Nature Publishing Group
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Document type: Journal Article
Collection: Office of the Deputy Vice-Chancellor Research
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