International Journal of Molecular Sciences | |
Validation of Reference Genes for the Determination of Platelet Transcript Level in Healthy Individuals and in Patients with the History of Myocardial Infarction | |
Katalin S. Zsóri2  László Muszbek2  Zoltán Csiki1  | |
[1] Department of Internal Medicine, University of Debrecen, Medical and Health Science Center, Debrecen 4032, Hungary; E-Mail:;Clinical Research Center, University of Debrecen, Medical and Health Science Center, Debrecen 4032, Hungary; E-Mails: | |
关键词: transcript level; normalization; platelet; reference gene; RT-qPCR; | |
DOI : 10.3390/ijms14023456 | |
来源: mdpi | |
【 摘 要 】
RT-qPCR is the standard method for studying changes in relative transcript level in different experimental and clinical conditions and in different tissues. No validated reference genes have been reported for the normalization of transcript level in platelets. The very low level of platelet RNA and the elimination of leukocyte contamination represented special methodological difficulties. Our aims were to apply a simple technique to separate platelets for transcript level studies, and select the most stable reference genes for platelets from healthy individuals and from patients with the history of myocardial infarction. We developed a simple, straightforward method of platelet separation for RNA isolation. Platelet activation was inhibited by using acid-citrate-dextrose for anticoagulation and by prostaglandin E1. Leukocyte contamination was eliminated by three consecutive centrifugations. Samples prepared by this method were free of leukocytes, showed no inhibition in PCR reaction and no RNA degradation. The assay demands low blood volume, which complies with the requirements of everyday laboratory routine. Seventeen potential reference genes were investigated, but eight of them were excluded during optimization. The stability of the remaining genes,
【 授权许可】
CC BY
© 2013 by the authors; licensee Molecular Diversity Preservation International, Basel, Switzerland.
【 预 览 】
Files | Size | Format | View |
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RO202003190038560ZK.pdf | 181KB | download |