Identification of endogenous reference genes for qRT-PCR analysis in normal matched breast tumor tissues

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dc.citation.epage365en_US
dc.citation.issueNumber8en_US
dc.citation.spage353en_US
dc.citation.volumeNumber17en_US
dc.contributor.authorGur-Dedeoglu, B.en_US
dc.contributor.authorKonu, O.en_US
dc.contributor.authorBozkurt, B.en_US
dc.contributor.authorErgul, G.en_US
dc.contributor.authorSeckin, S.en_US
dc.contributor.authorYulug, I. G.en_US
dc.date.accessioned2016-02-08T10:01:00Z
dc.date.available2016-02-08T10:01:00Z
dc.date.issued2009en_US
dc.departmentDepartment of Molecular Biology and Geneticsen_US
dc.description.abstractQuantitative gene expression measurements from tumor tissue are frequently compared with matched normal and/or adjacent tumor tissue expression for diagnostic marker gene selection as well as assessment of the degree of transcriptional deregulation in cancer. Selection of an appropriate reference gene (RG) or an RG panel, which varies depending on cancer type, molecular subtypes, and the normal tissues used for interindividual calibration, is crucial for the accurate quantification of gene expression. Several RG panels have been suggested in breast cancer for making comparisons among tumor subtypes, cell lines, and benign/malignant tumors. In this study, expression patterns of 15 widely used endogenous RGs (ACTB, TBP, GAPDH, SDHA, HPRT, HMBS, B2M, PPIA, GUSB, YWHAZ2, PGK1, RPLP0, PUM1, MRPL19, and RPL41), and three candidate genes that were selected through analysis of two independent microarray datasets (IL22RA1, TTC22, ZNF224) were determined in 23 primary breast tumors and their matched normal tissues using qRTPCR. Additionally, 18S rRNA, ACTB, and SDHA were tested using randomly primed cDNAs from 13 breast tumor pairs to assess the rRNA/mRNA ratio. The tumors exhibited significantly lower rRNA/mRNA ratio when compared to their normals, on average. The expression of the studied RGs in breast tumors did not exhibit differences in terms of grade, ER, or PR status. The stability of RGs was examined based on two different statistical models, namely GeNorm and NormFinder. Among the 18 tested endogenous reference genes, ACTB and SDHA were identified as the most suitable reference genes for the normalization of qRTPCR data in the analysis of normal matched tumor breast tissue pairs by both programs. In addition, the expression of the gelsolin (GSN) gene, a well-known downregulated target in breast tumors, was analyzed using the two most suitable genes and different RG combinations to validate their effectiveness as a normalization factor (NF). The GSN expression of the tumors used in this study was significantly lower than that of normals showing the effectivity of using ACTB and SDHA as suitable RGs in this set of tumor–normal tissue panel. The combinational use of the best performing two RGs (ACTB and SDHA) as a normalization factor can be recommended to minimize sample variability and to increase the accuracy and resolution of gene expression normalization in tumor–normal paired breast cancer qRT-PCR studies.en_US
dc.description.provenanceMade available in DSpace on 2016-02-08T10:01:00Z (GMT). No. of bitstreams: 1 bilkent-research-paper.pdf: 70227 bytes, checksum: 26e812c6f5156f83f0e77b261a471b5a (MD5) Previous issue date: 2009en
dc.identifier.doi10.3727/096504009788428460en_US
dc.identifier.issn1555-3906
dc.identifier.urihttp://hdl.handle.net/11693/22506
dc.language.isoEnglishen_US
dc.publisherCognizant Communication Corporationen_US
dc.relation.isversionof10.3727/096504009788428460en_US
dc.source.titleOncology Researchen_US
dc.subjectBreast canceren_US
dc.subjectEndogenous reference genesen_US
dc.subjectNormalization factoren_US
dc.subjectReal-time quantitative RT-PCRen_US
dc.subjectbeta 2 microglobulinen_US
dc.subjectbeta actinen_US
dc.subjectbeta glucuronidaseen_US
dc.subjectcyclophilin Aen_US
dc.subjectDrosophila proteinen_US
dc.subjectgelsolinen_US
dc.subjectglyceraldehyde 3 phosphate dehydrogenaseen_US
dc.subjecthypoxanthine phosphoribosyltransferaseen_US
dc.subjectinterleukin 22en_US
dc.subjectinterleukin 22 receptor alpha 1en_US
dc.subjectmessenger RNAen_US
dc.subjectmitochondrial ribosomal protein L19en_US
dc.subjectphosphoglycerate kinaseen_US
dc.subjectpolypeptideen_US
dc.subjectporphobilinogen deaminaseen_US
dc.subjectPumilio homolog 1en_US
dc.subjectribosomal protein L41en_US
dc.subjectribosomal protein large P0en_US
dc.subjectribosome proteinen_US
dc.subjectribosome RNAen_US
dc.subjectRNA 18Sen_US
dc.subjectsuccinate dehydrogenaseen_US
dc.subjectsuccinate dehydrogenase complex subunit Aen_US
dc.subjectTATA binding proteinen_US
dc.subjecttetratricopeptide repeat domain 22en_US
dc.subjecttetratricopeptide repeat proteinen_US
dc.subjecttyrosine 3 monooxygenase tryptophan 5 monooxygenase activation protein zeta polypeptideen_US
dc.subjectunclassified drugen_US
dc.subjectzinc finger proteinen_US
dc.subjectzinc finger protein 224en_US
dc.subjecttumor markeren_US
dc.subjectadulten_US
dc.subjectageden_US
dc.subjectarticleen_US
dc.subjectbase pairingen_US
dc.subjectcancer gradingen_US
dc.subjectclinical articleen_US
dc.subjectcontrolled studyen_US
dc.subjectDNA microarrayen_US
dc.subjectdown regulationen_US
dc.subjectgene expressionen_US
dc.subjectgene targetingen_US
dc.subjectgenetic analysisen_US
dc.subjectgenetic variabilityen_US
dc.subjectgenomic instabilityen_US
dc.subjecthumanen_US
dc.subjecthuman tissueen_US
dc.subjectmolecular cloningen_US
dc.subjectnucleotide sequenceen_US
dc.subjectpriority journalen_US
dc.subjectreverse transcription polymerase chain reactionen_US
dc.subjectbreast tumoren_US
dc.subjectfemaleen_US
dc.subjectgene expression profilingen_US
dc.subjectgeneticsen_US
dc.subjectreverse transcription polymerase chain reactionen_US
dc.subjectstandarden_US
dc.subjectBreast Neoplasmsen_US
dc.subjectFemaleen_US
dc.subjectGene Expression Profilingen_US
dc.subjectHumansen_US
dc.subjectReference Standardsen_US
dc.subjectReverse Transcriptase Polymerase Chain Reactionen_US
dc.subjectTumor Markers, Biologicalen_US
dc.titleIdentification of endogenous reference genes for qRT-PCR analysis in normal matched breast tumor tissuesen_US
dc.typeArticleen_US

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