Identification and validation of suitable endogenous reference genes for gene expression studies in human peripheral blood

Stamova, Boryana; Apperson, Michelle; Walker, Wynn L.; Tian, Yingfang; Xu, Huichun; Adamczy, Peter; Zhan, Xinhua; Liu, Dazhi; Ander, Bradley P.; Liao, Isaac; Gregg, Jeffrey P.; Turner, Renée J.; Jickling, Glen C.; Lit, Lisa; Sharp, Frank R.

doi:10.1186/1755-8794-2-49

Cited by 100 publications

(91 citation statements)

References 39 publications

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“…However, there are several limitations and potential confounding factors. For example, although we have found that GAPDH mRNA expression in blood is stable in previous studies , replication of these biomarkers using several reference genes for normalization is desirable (Stamova, et al, 2009). In addition, differences in blood counts and PD medications may bias gene expression results.…”

Section: Discussionmentioning

confidence: 99%

Splice Variant Biomarkers for Parkinson's Disease

Potashkin¹

2014

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Public reporting burden for this collection of information is estimated to average 1 hour per response, including the time for reviewing instructions, searching existing data sources, gathering and maintaining the data needed, and completing and reviewing this collection of information. Send comments regarding this burden estimate or any other aspect of this collection of information, including suggestions for reducing this burden to Department of Defense, Washington Headquarters Services, Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson Davis Highway, Suite 1204, Arlington, VA 22202-4302. Respondents should be aware that notwithstanding any other provision of law, no person shall be subject to any penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number. PLEASE DO NOT RETURN YOUR FORM TO THE ABOVE ADDRESS. REPORT DATEMay PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) AND ADDRESS(ES) PERFORMING ORGANIZATION REPORT NUMBERRosalind Franklin U. Medicine & Science 3333 Green Bay Rd. North Chicago, IL 60064-3037 SPONSORING / MONITORING AGENCY NAME(S) AND ADDRESS(ES) 10. SPONSOR/MONITOR'S ACRONYM(S) U.S. Army Medical Research and Materiel Command Fort Detrick, Maryland 21702-5012 SPONSOR/MONITOR'S REPORT NUMBER(S) DISTRIBUTION / AVAILABILITY STATEMENTApproved for Public Release; Distribution Unlimited SUPPLEMENTARY NOTES ABSTRACTRecently we identified splice variants in whole blood than can distinguish early stage Parkinson's disease (PD) patients from healthy and neurodegenerative controls. The proposed studies will test the hypothesis that the biomarkers will be useful for identifying individuals at risk for PD and for identifying signaling pathways that are disrupted in PD. We are testing the expression of the biomarkers in RNA prepared from whole blood of hyposmic participants in the Parkinson's Associated Risk Study (PARS) (Technical Objective 1.0). In order to establish a model for testing the function of the biomarkers, we are determining whether their expression is altered in human olfactory neurosphere-derived (hONS) cells derived from idiopathic PD patients and healthy controls (Technical Objective 2.0). To determine the signaling pathways affected in PD and identify additional biomarkers, we are using network analysis (Technical Objective 3.0). During year 1, RNA and cDNA was prepared from PARS samples from Year 0 (baseline) and Year 2. Quantitative polymerase chain reactions were initiated for two of the biomarkers (APP, HNF4α). Expression of the biomarkers was tested in hONS. We also developed network approaches to reveal the common molecular pathways involved with PD and type 2 diabetes (T2DM). Using these networks, we identified APP, HNF4A and SOD2 mRNAs as blood biomarkers predictive of early stage PD. In addition, we determined that HNF4A mRNA may be a progression marker in blood for PD. SUBJECT TERMSParkinson's disease, biomarkers, neurodegeneration, network and pathway analysis 12-15Santiago 16-23Sei...

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Section: Discussionmentioning

confidence: 99%

Splice Variant Biomarkers for Parkinson's Disease

Potashkin¹

2014

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“…However, other studies have shown that these too can be affected by various experimental conditions and requires prior validation (Bionaz and Loor, 2007;Bustin, 2000;Chapman and Waldenström, 2015;Glare et al, 2002;Stamova et al, 2009). The present study found GAPDH to be a suitable endogenous in the SM and LT of young bulls, SM of steers and the LT of heifers.…”

Section: Discussionmentioning

confidence: 99%

Expression stability of reference genes in the skeletal muscles of beef cattle

Mberema¹,

Sparagano²

2017

Afr. J. Biotechnol.

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The objectives of this study were to evaluate the relative expression stability of five candidate reference genes in the semimembranosus (SM) and longissimus thoracis (LT) of beef steers, heifers and young bulls. The mRNA levels of Beta-Actin, eukaryotic initiation factor-2B Subunit 2, glyceraldehyde-3-phosphate dehydrogenase, peptidylprolyl isomerase-A and succinate dehydrogenase complex-subunit A were quantified using real-time polymerase chain reaction (qPCR). The combined analysis using the geNorm algorithm revealed that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and eukaryotic initiation factor-2B subunit 2 (EIF2B2) were the most stable gene pairs. However, individual experimental conditions showed that succinate dehydrogenase complex-subunit A was most stably expressed in bulls and heifers SM, and in bulls and steers LT. Glyceraldehyde-3-phosphate dehydrogenase was most stably expressed in bull SM, steer SM, bull LT and heifer LT. The expression stability ranking order differed between experimental conditions, but all genes had low expression variability. Therefore, using the two most stable reference genes, namely GAPDH and EIF2B2, would result in more accurate normalizations for quantitative real-time PCR studies in the SM and LT muscles of beef cattle. The need for prior evaluation of candidate reference genes in different muscles and sex groups of beef cattle is thus emphasized by the present results.

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“…Six candidate reference genes (ACTB, CRY2, CSNK1G2, DECR1, FARP1, and TRAP1) that have been reported to be consistently expressed in human whole-blood samples were selected for investigation (11). The expression stability of the 6 reference genes in the training set were estimated using 4 commonly used algorithms: geNorm (12), NormFinder (13), BestKeeper (14), and the comparative cycle threshold (DC t ) method (15).…”

Section: Qrt-pcrmentioning

confidence: 99%

Identification and Validation of a Blood-Based 18-Gene Expression Signature in Colorectal Cancer

et al. 2013

Clinical Cancer Research

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Purpose: The early detection of colorectal cancer (CRC) is crucial for successful treatment and patient survival. However, compliance with current screening methods remains poor. This study aimed to identify an accurate blood-based gene expression signature for CRC detection.Experimental Design: Gene expression in peripheral blood samples from 216 patients with CRC tumors and 187 controls was investigated in the study. We first conducted a microarray analysis to select candidate genes that were significantly differentially expressed between patients with cancer and controls. A quantitative reverse transcription PCR assay was then used to evaluate the expression of selected genes. A gene expression signature was identified using a training set (n ¼ 200) and then validated using an independent test set (n ¼ 160).Results: We identified an 18-gene signature that discriminated the patients with CRC from controls with 92% accuracy, 91% sensitivity, and 92% specificity. The signature performance was further validated in the independent test set with 86% accuracy, 84% sensitivity, and 88% specificity. The area under the receiver operating characteristics curve was 0.94. The signature was shown to be enriched in genes related to immune functions.Conclusions: This study identified an 18-gene signature that accurately discriminated patients with CRC from controls in peripheral blood samples. Our results prompt the further development of blood-based gene expression biomarkers for the diagnosis and early detection of CRC.

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Identification and validation of suitable endogenous reference genes for gene expression studies in human peripheral blood

Cited by 100 publications

References 39 publications

Splice Variant Biomarkers for Parkinson's Disease

Splice Variant Biomarkers for Parkinson's Disease

Expression stability of reference genes in the skeletal muscles of beef cattle

Identification and Validation of a Blood-Based 18-Gene Expression Signature in Colorectal Cancer

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