title: Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co γ-rays ex vivo.
abstract: In gene expression studies, it is critical to normalize data using a stably expressed endogenous control gene in order to obtain accurate and reliable results. However, we currently do not have a universally applied endogenous control gene for normalization of data for gene expression studies, particularly those involving (60)Co γ-ray-exposed human blood samples. In this study, a comparative assessment of the gene expression of six widely used housekeeping endogenous control genes, namely 18S, ACTB, B2M, GAPDH, MT-ATP6 and CDKN1A, was undertaken for a range of (60)Co γ-ray doses (0.5, 1.0, 2.0 and 4.0 Gy) at 8.4 Gy min(-1) at 0 and 24 h post-irradiation time intervals. Using the NormFinder algorithm, real-time PCR data obtained from six individuals (three males and three females) were analyzed with respect to the threshold cycle (Ct) value and abundance, ΔCt pair-wise comparison, intra- and inter-group variability assessments, etc. GAPDH, either alone or in combination with 18S, was found to be the most suitable endogenous control gene and should be used in gene expression studies, especially those involving qPCR of γ-ray-exposed human blood samples.
authors.affiliation: Radiation and Molecular Biology Unit, Department of Biochemistry, North-Eastern Hill University (NEHU), Shillong, 793022, India.
authors.forename: S Thangminlal
authors.initials: ST
authors.lastname: Vaiphei
authors.affiliation: Radiation and Molecular Biology Unit, Department of Biochemistry, North-Eastern Hill University (NEHU), Shillong, 793022, India.
authors.forename: Joshua
authors.initials: J
authors.lastname: Keppen
authors.affiliation: Radiation and Molecular Biology Unit, Department of Biochemistry, North-Eastern Hill University (NEHU), Shillong, 793022, India.
authors.forename: Saibadaiahun
authors.initials: S
authors.lastname: Nongrum
authors.affiliation: Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre (BARC), Trombay, Mumbai, 400085, India.
authors.forename: R C
authors.initials: RC
authors.lastname: Chaubey
authors.affiliation: Radiation Countermeasures Unit, Department of Biochemistry, North-Eastern Hill University (NEHU), Shillong, 793022, India.
authors.forename: L
authors.initials: L
authors.lastname: Kma
authors.affiliation: Radiation and Molecular Biology Unit, Department of Biochemistry, North-Eastern Hill University (NEHU), Shillong, 793022, India rnsharan@nehu.ac.in.
authors.forename: R N
authors.initials: RN
authors.lastname: Sharan
mesh_terms: D000328:Adult; D001681:Biological Assay / Q000592:standards*; D001769:Blood / Q000378:metabolism* / Q000528:radiation effects*; D001798:Blood Proteins / Q000378:metabolism*; D003037:Cobalt Radioisotopes; D004307:Dose-Response Relationship, Radiation; D005260:Female; D005720:Gamma Rays; D020869:Gene Expression Profiling / Q000592:standards; D005786:Gene Expression Regulation / Q000502:physiology* / Q000528:radiation effects; D006801:Humans; D007194:India; D008297:Male; D011829:Radiation Dosage; D012016:Reference Values; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D001798:Blood Proteins; D003037:Cobalt Radioisotopes
keywords: 60Co γ-rays; endogenous control gene; gene expression; human peripheral blood lymphocyte (HPBL); qPCR