Design, optimization and validation of genes commonly used in expression studies on DMH/AOM rat colon carcinogenesis model
| Autor: | Bars-Cortina, David, Riera-Escamilla, Antoni, Gou Alsina, Gemma, Piñol-Felis, Carme, Motilva, María-José, Universitat Autònoma de Barcelona |
|---|---|
| Přispěvatelé: | Ministerio de Educación, Cultura y Deporte (España) |
| Rok vydání: | 2019 |
| Předmět: |
Colorectal cancer
Colon Metabolite SYBR Azoxymethane lcsh:Medicine Biology General Biochemistry Genetics and Molecular Biology Melting curve analysis 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Reference genes medicine Dimethylhydrazine (DMH Molecular Biology Gene Nutrition 030304 developmental biology Azoxymethane (AOM) 0303 health sciences General Neuroscience lcsh:R Cancer General Medicine Dimethylhydrazine Food Science and Technology medicine.disease QPCR Gene validation qPCR Oncology chemistry 030220 oncology & carcinogenesis Cancer research Primer (molecular biology) General Agricultural and Biological Sciences Dimethylhydrazine (DMH) |
| Zdroj: | Dipòsit Digital de Documents de la UAB Universitat Autònoma de Barcelona RIUR. Repositorio Institucional de la Universidad de La Rioja instname Repositorio Abierto de la UdL Universitad de Lleida Recercat. Dipósit de la Recerca de Catalunya Digital.CSIC. Repositorio Institucional del CSIC PeerJ, Vol 7, p e6372 (2019) PeerJ r-IIB SANT PAU. Repositorio Institucional de Producción Científica del Instituto de Investigación Biomédica Sant Pau |
| ISSN: | 2167-8359 |
| Popis: | Colorectal cancer (CRC), also known as colon cancer, is the third most common form of cancer worldwide in men and the second in women and is characterized by several genetic alterations, among them the expression of several genes. 1,2-dimethylhydrazine (DMH) and its metabolite azoxymethane (AOM) are procarcinogens commonly used to induce colon cancer in rats (DMH/AOM rat model). This rat model has been used to study changes in mRNA expression in genes involved in this pathological condition. However, a lack of proper detailed PCR primer design in the literature limits the reproducibility of the published data. The present study aims to design, optimize and validate the qPCR, in accordance with the MIQE (Minimum Information for Publication of Quantitative Real-Time PCR Experiments) guidelines, for seventeen genes commonly used in the DMH/AOM rat model of CRC (Apc, Aurka, Bax, Bcl2, β-catenin, Ccnd1, Cdkn1a, Cox2, Gsk3beta, IL-33, iNOs, Nrf2, p53, RelA, Smad4, Tnf α and Vegfa) and two reference genes (Actb or β-actin and B2m). The specificity of all primer pairs was empirically validated on agarose gel, and furthermore, the melting curve inspection was checked as was their efficiency (%) ranging from 90 to 110 with a correlation coefficient of r 2 > 0.980. Finally, a pilot study was performed to compare the robustness of two candidate reference genes. This study was supported by the Spanish Ministry of Education, Culture and Sport through the ‘‘Formación Profesorado Universitario (FPU)’’ grant awarded to David Bars-Cortina (FPU014/02902) |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|