• Login
    View Item 
    •   Home
    • The Christie Research Publications Repository
    • All Christie Publications
    • View Item
    •   Home
    • The Christie Research Publications Repository
    • All Christie Publications
    • View Item
    JavaScript is disabled for your browser. Some features of this site may not work without it.

    Browse

    All of ChristieCommunitiesTitleAuthorsIssue DateSubmit DateSubjectsThis CollectionTitleAuthorsIssue DateSubmit DateSubjectsProfilesView

    My Account

    LoginRegister

    Local Links

    The Christie WebsiteChristie Library and Knowledge Service

    Statistics

    Display statistics

    Selection of endogenous control genes for normalising gene expression data derived from formalin-fixed paraffin-embedded tumour tissue

    • CSV
    • RefMan
    • EndNote
    • BibTex
    • RefWorks
    Thumbnail
    Name:
    33057113.pdf
    Size:
    1.143Mb
    Format:
    PDF
    Description:
    From UNPAYWALL
    Download
    Authors
    Smith, Tim A D
    AbdelKarem, Omneya A
    Irlam-Jones, Joely J
    Lane, Brian
    Valentine, Helen R
    Bibby, Becky A
    Denley, H.
    Choudhury, Ananya
    West, Catharine M L
    Affiliation
    Translational Radiobiology Group, Division of Cancer Sciences, University of Manchester, Manchester Academic Health Centre, Christie Hospital NHS Found Trust, Manchester, M20 4BX, UK.
    Issue Date
    2020
    
    Metadata
    Show full item record
    Abstract
    Quantitative real time polymerase chain reaction (qPCR) data are normalised using endogenous control genes. We aimed to: (1) demonstrate a pathway to identify endogenous control genes for qPCR analysis of formalin-fixed paraffin-embedded (FFPE) tissue using bladder cancer as an exemplar; and (2) examine the influence of probe length and sample age on PCR amplification and co-expression of candidate genes on apparent expression stability. RNA was extracted from prospective and retrospective samples and subject to qPCR using TaqMan human endogenous control arrays or single tube assays. Gene stability ranking was assessed using coefficient of variation (CoV), GeNorm and NormFinder. Co-expressed genes were identified from The Cancer Genome Atlas (TCGA) using the on-line gene regression analysis tool GRACE. Cycle threshold (Ct) values were lower for prospective (19.49 ± 2.53) vs retrospective (23.8 ± 3.32) tissues (p < 0.001) and shorter vs longer probes. Co-expressed genes ranked as the most stable genes in the TCGA cohort by GeNorm when analysed together but ranked lower when analysed individually omitting co-expressed genes indicating bias. Stability values were < 1.5 for the 20 candidate genes in the prospective cohort. As they consistently ranked in the top ten by CoV, GeNorm and Normfinder, UBC, RPLP0, HMBS, GUSB, and TBP are the most suitable endogenous control genes for bladder cancer qPCR.
    Citation
    Smith TAD, AbdelKarem OA, Irlam-Jones JJ, Lane B, Valentine H, Bibby BAS, et al. Selection of endogenous control genes for normalising gene expression data derived from formalin-fixed paraffin-embedded tumour tissue. Sci Rep. 2020;10(1):17258.
    Journal
    Scientific Reports
    URI
    http://hdl.handle.net/10541/623435
    DOI
    10.1038/s41598-020-74380-7
    PubMed ID
    33057113
    Additional Links
    https://dx.doi.org/10.1038/s41598-020-74380-7
    Type
    Article
    Language
    en
    ae974a485f413a2113503eed53cd6c53
    10.1038/s41598-020-74380-7
    Scopus Count
    Collections
    All Christie Publications

    entitlement

    Related articles

    • Identification of accurate reference genes for RT-qPCR analysis of formalin-fixed paraffin-embedded tissue from primary non-small cell lung cancers and brain and lymph node metastases.
    • Authors: Søes S, Sørensen BS, Alsner J, Overgaard J, Hager H, Hansen LL, Kristensen LS
    • Issue date: 2013 Aug
    • The use of laser microdissection in the identification of suitable reference genes for normalization of quantitative real-time PCR in human FFPE epithelial ovarian tissue samples.
    • Authors: Cai J, Li T, Huang B, Cheng H, Ding H, Dong W, Xiao M, Liu L, Wang Z
    • Issue date: 2014
    • Development and evaluation of a novel RT-qPCR based test for the quantification of HER2 gene expression in breast cancer.
    • Authors: El Hadi H, Abdellaoui-Maane I, Kottwitz D, El Amrani M, Bouchoutrouch N, Qmichou Z, Karkouri M, ElAttar H, Errihani H, Fernandez PL, Bakri Y, Sefrioui H, Moumen A
    • Issue date: 2017 Mar 20
    • Validation of endogenous control genes for gene expression studies on human ocular surface epithelium.
    • Authors: Kulkarni B, Mohammed I, Hopkinson A, Dua HS
    • Issue date: 2011
    • Validation of reference genes for quantitative real-time PCR studies in the dentate gyrus after experimental febrile seizures.
    • Authors: Swijsen A, Nelissen K, Janssen D, Rigo JM, Hoogland G
    • Issue date: 2012 Dec 13
    DSpace software (copyright © 2002 - 2025)  DuraSpace
    Quick Guide | Contact Us
    Open Repository is a service operated by 
    Atmire NV
     

    Export search results

    The export option will allow you to export the current search results of the entered query to a file. Different formats are available for download. To export the items, click on the button corresponding with the preferred download format.

    By default, clicking on the export buttons will result in a download of the allowed maximum amount of items.

    To select a subset of the search results, click "Selective Export" button and make a selection of the items you want to export. The amount of items that can be exported at once is similarly restricted as the full export.

    After making a selection, click one of the export format buttons. The amount of items that will be exported is indicated in the bubble next to export format.