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Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co ?-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 ?-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 ?-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, ?Ct 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 ?-ray-exposed human blood samples.

SUBMITTER: Vaiphei ST

PROVIDER: S-EPMC4572586 | biostudies-literature | 2015 Jan

REPOSITORIES: biostudies-literature

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Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co γ-rays ex vivo.

Vaiphei S Thangminlal ST Keppen Joshua J Nongrum Saibadaiahun S Chaubey R C RC Kma L L Sharan R N RN

Journal of radiation research 20140930 1

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, A ...[more]

PMID: 25271263

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Project description:In the current global scenario there is always a possibility that a terrorist group might acquire material to produce a nuclear device. In the likelihood of a detonation of an improvised nuclear device (IND) the prompt radiation would consist of both photons (gamma rays) and neutrons. As neutrons generally have a high Relative Biological Effectiveness (RBE) for most physiological endpoints, it is important to understand the impact that neutrons would have on the biodosimetry methods that are being developed for medical triage purposes. We have previously reported transcriptomic response in human and mouse blood to neutron exposure and determined its RBE compared to photons for gene induction. In this report, we studied the effect of mixed neutron-photon radiation on gene expression in human peripheral blood in order to mimic an IND type radiation observed increase in number as well as expression level of genes with increasing percentage of neutron in mixed exposures which peaked at 25% neutron exposure which was maximum neutron percentage used for the study. We identified 156 genes that responded significantly to all the neutron-photon mixed exposures. These genes were found to be mainly involved in p53 signaling and DNA damage response. The 25% neutron exposure also showed maximum enrichment of known neutron associated gene ontology terms reported in the previous studies. Analysis of upstream regulators of gene expression showed strong activation of TP53, SLC29A1, PDCD1 and suppression of ANLN, AURK and ANXA2 across all exposures. While AGT was exclusively activated by neutron specific exposures. We confirmed the expression of genes which showed increased expression as a function of increased neutron percentage using quantitative real-time RT-PCR which matched with the microarray data. Thus, the data in this current study clearly indicates that gene expression can be used to estimate the percentage of the neutron component in mixed neutron-photon exposures.

Dataset Information

Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co ?-rays ex vivo.

Publications

Evaluation of endogenous control gene(s) for gene expression studies in human blood exposed to 60Co γ-rays ex vivo.

Similar Datasets

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