TY - JOUR
T1 - An international comparability study on quantification of mRNA gene expression ratios
T2 - CCQM-P103.1
AU - Devonshire, Alison S.
AU - Sanders, Rebecca
AU - Whale, Alexandra S.
AU - Nixon, Gavin J.
AU - Cowen, Simon
AU - Ellison, Stephen L.R.
AU - Parkes, Helen
AU - Pine, P. Scott
AU - Salit, Marc
AU - McDaniel, Jennifer
AU - Munro, Sarah
AU - Lund, Steve
AU - Matsukura, Satoko
AU - Sekiguchi, Yuji
AU - Kawaharasaki, Mamoru
AU - Granjeiro, José Mauro
AU - Falagan-Lotsch, Priscila
AU - Saraiva, Antonio Marcos
AU - Couto, Paulo
AU - Yang, Inchul
AU - Kwon, Hyerim
AU - Park, Sang Ryoul
AU - Demšar, Tina
AU - Žel, Jana
AU - Blejec, Andrej
AU - Milavec, Mojca
AU - Dong, Lianhua
AU - Zhang, Ling
AU - Sui, Zhiwei
AU - Wang, Jing
AU - Viroonudomphol, Duangkamol
AU - Prawettongsopon, Chaiwat
AU - Partis, Lina
AU - Baoutina, Anna
AU - Emslie, Kerry
AU - Takatsu, Akiko
AU - Akyurek, Sema
AU - Akgoz, Muslum
AU - Vonsky, Maxim
AU - Konopelko, L. A.
AU - Cundapi, Edna Matus
AU - Urquiza, Melina Pérez
AU - Huggett, Jim F.
AU - Foy, Carole A.
N1 - Funding Information:
The work described in this paper was funded in part by the UK National Measurement System .
Publisher Copyright:
© 2016.
PY - 2016/6/1
Y1 - 2016/6/1
N2 - Measurement of RNA can be used to study and monitor a range of infectious and non-communicable diseases, with profiling of multiple gene expression mRNA transcripts being increasingly applied to cancer stratification and prognosis. An international comparison study (Consultative Committee for Amount of Substance (CCQM)-P103.1) was performed in order to evaluate the comparability of measurements of RNA copy number ratio for multiple gene targets between two samples. Six exogenous synthetic targets comprising of External RNA Control Consortium (ERCC) standards were measured alongside transcripts for three endogenous gene targets present in the background of human cell line RNA. The study was carried out under the auspices of the Nucleic Acids (formerly Bioanalysis) Working Group of the CCQM. It was coordinated by LGC (United Kingdom) with the support of National Institute of Standards and Technology (USA) and results were submitted from thirteen National Metrology Institutes and Designated Institutes. The majority of laboratories performed RNA measurements using RT-qPCR, with datasets also being submitted by two laboratories based on reverse transcription digital polymerase chain reaction and one laboratory using a next-generation sequencing method. In RT-qPCR analysis, the RNA copy number ratios between the two samples were quantified using either a standard curve or a relative quantification approach. In general, good agreement was observed between the reported results of ERCC RNA copy number ratio measurements. Measurements of the RNA copy number ratios for endogenous genes between the two samples were also consistent between the majority of laboratories. Some differences in the reported values and confidence intervals ('measurement uncertainties') were noted which may be attributable to choice of measurement method or quantification approach. This highlights the need for standardised practices for the calculation of fold change ratios and uncertainties in the area of gene expression profiling.
AB - Measurement of RNA can be used to study and monitor a range of infectious and non-communicable diseases, with profiling of multiple gene expression mRNA transcripts being increasingly applied to cancer stratification and prognosis. An international comparison study (Consultative Committee for Amount of Substance (CCQM)-P103.1) was performed in order to evaluate the comparability of measurements of RNA copy number ratio for multiple gene targets between two samples. Six exogenous synthetic targets comprising of External RNA Control Consortium (ERCC) standards were measured alongside transcripts for three endogenous gene targets present in the background of human cell line RNA. The study was carried out under the auspices of the Nucleic Acids (formerly Bioanalysis) Working Group of the CCQM. It was coordinated by LGC (United Kingdom) with the support of National Institute of Standards and Technology (USA) and results were submitted from thirteen National Metrology Institutes and Designated Institutes. The majority of laboratories performed RNA measurements using RT-qPCR, with datasets also being submitted by two laboratories based on reverse transcription digital polymerase chain reaction and one laboratory using a next-generation sequencing method. In RT-qPCR analysis, the RNA copy number ratios between the two samples were quantified using either a standard curve or a relative quantification approach. In general, good agreement was observed between the reported results of ERCC RNA copy number ratio measurements. Measurements of the RNA copy number ratios for endogenous genes between the two samples were also consistent between the majority of laboratories. Some differences in the reported values and confidence intervals ('measurement uncertainties') were noted which may be attributable to choice of measurement method or quantification approach. This highlights the need for standardised practices for the calculation of fold change ratios and uncertainties in the area of gene expression profiling.
KW - Biomarker identification and validation
KW - Cancer
KW - Diagnostics
KW - Gene expression
KW - Molecular diagnostic
KW - Normalisation
KW - RNA copy number ratio
KW - RT-qPCR
KW - Standardisation
KW - Transcriptomics
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U2 - 10.1016/j.bdq.2016.05.003
DO - 10.1016/j.bdq.2016.05.003
M3 - Article
AN - SCOPUS:84973137565
SN - 2214-7535
VL - 8
SP - 15
EP - 28
JO - Biomolecular Detection and Quantification
JF - Biomolecular Detection and Quantification
ER -
