Dataset Information

Selection of suitable reference genes for normalization of quantitative RT-PCR in peripheral blood samples of bottlenose dolphins (Tursiops truncatus).

ABSTRACT: Quantitative RT-PCR is often used as a research tool directed at gene transcription. Selection of optimal housekeeping genes (HKGs) as reference genes is critical to establishing sensitive and reproducible qRT-PCR-based assays. The current study was designed to identify the appropriate reference genes in blood leukocytes of bottlenose dolphins (Tursiops truncatus) for gene transcription research. Seventy-five blood samples collected from 7 bottlenose dolphins were used to analyze 15 candidate HKGs (ACTB, B2M, GAPDH, HPRT1, LDHB, PGK1, RPL4, RPL8, RPL18, RPS9, RPS18, TFRC, YWHAZ, LDHA, SDHA). HKG stability in qRT-PCR was determined using geNorm, NormFinder, BestKeeper and comparative delta Ct algorithms. Utilization of RefFinder, which combined all 4 algorithms, suggested that PGK1, HPRT1 and RPL4 were the most stable HKGs in bottlenose dolphin blood. Gene transcription perturbations in blood can serve as an indication of health status in cetaceans as it occurs prior to alterations in hematology and chemistry. This study identified HKGs that could be used in gene transcript studies, which may contribute to further mRNA relative quantification research in the peripheral blood leukocytes in captive cetaceans.

SUBMITTER: Chen IH

PROVIDER: S-EPMC4614023 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Publications

Selection of suitable reference genes for normalization of quantitative RT-PCR in peripheral blood samples of bottlenose dolphins (Tursiops truncatus).

Chen I-Hua IH Chou Lien-Siang LS Chou Shih-Jen SJ Wang Jiann-Hsiung JH Stott Jeffrey J Blanchard Myra M Jen I-Fan IF Yang Wei-Cheng WC

Scientific reports 20151021

Quantitative RT-PCR is often used as a research tool directed at gene transcription. Selection of optimal housekeeping genes (HKGs) as reference genes is critical to establishing sensitive and reproducible qRT-PCR-based assays. The current study was designed to identify the appropriate reference genes in blood leukocytes of bottlenose dolphins (Tursiops truncatus) for gene transcription research. Seventy-five blood samples collected from 7 bottlenose dolphins were used to analyze 15 candidate HK ...[more]

PMID: 26486099

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Project description:As apex predators, cetaceans are recognized as sentinels for the health of marine ecosystems. The ability to identify the impacts of anthropogenic contamination on the health of a species requires long term monitoring to establish baseline health parameters. This makes health measurements that can be sampled from free ranging animals highly desirable. In the current study, we investigate the utility of skin gene expression profiling to monitor the physiological, health, and contaminant exposure status in bottlenose dolphins. Remote integument biopsies were obtained from bottlenose dolphins at three locations in the northern Gulf of Mexico prior to oil exposure (May 2010) and during summer and winter of the two years (2010-2011) following their exposure to oil from the Deepwater Horizon oil spill. A bottlenose dolphin-specific oligonucleotide microarray was used to characterize the skin transcriptomes of 94 individuals from three genetically distinguishable populations in Barataria Bay, Louisiana, Chandeleur Sound, Louisiana, and Mississippi Sound, Mississippi. Location did not significantly affect the transcriptome patterns of the dolphins included in the study. In contrast, season had a profound effect on gene expression, with nearly one-third of all genes on the array differing significantly in expression between winter and the warmer seasons (p<0.01, fold-change >1.5). Gene ontology enrichment analysis revealed that processes related to cell proliferation, motility and differentiation dominated the seasonal differences in expression, which likely reflects the differences in ambient temperature to which the skin is exposed, and possibly changes in blood flow. More subtle differences were seen between spring and summer transcriptomes (1.5% differentially expressed), with two presumed oil-exposed animals presenting gene expression profiles more similar to the summer (exposed) animals than the other spring animals. Xenobiotic pathway components also showed some seasonal trends, which may reflect crosstalk between xenobiotic pathways and those regulating corticoids, hormones, and cytokines. Seasonal effects have not previously been considered in studies assessing gene expression in cetaceans, but clearly must be taken into account when applying transcriptomic analyses to investigate their exposure or health status.

Dataset Information

Selection of suitable reference genes for normalization of quantitative RT-PCR in peripheral blood samples of bottlenose dolphins (Tursiops truncatus).

Publications

Selection of suitable reference genes for normalization of quantitative RT-PCR in peripheral blood samples of bottlenose dolphins (Tursiops truncatus).

Similar Datasets

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