1. Selection of Reference Genes for Quantitative Real Time PCR (qPCR) Assays in Tissue from Human Ascending Aorta
- Author
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Borja Fernández, Juan Robledo-Carmona, Carmen Rueda-Martínez, Gemma Sánchez-Espín, Manuel F. Jiménez-Navarro, Maria Mataró, Miguel Such-Martínez, Oscar Lamas, [Rueda-Martínez,C, Lamas,O, Mataró,MJ, Robledo-Carmona,J, Sánchez-Espín,G, Jiménez-Navarro,M, Such-Martínez,M] UGC del Corazón, Instituto de Investigación Biomédica de Málaga (IBIMA), Hospital Clínico Universitario Virgen de la Victoria, Málaga, Spain. [Jiménez-Navarro,M] Departamento de Medicina, Facultad de Medicina, Universidad de Málaga, Spain. [Fernández, B] Departamento de Biología Animal, Instituto de Investigación Biomédica de Málaga (IBIMA), Facultad de Ciencias, Universidad de Málaga, Spain., and This work was supported in part by grants PI-0689/2010 (Consejería de Salud, Junta de Andalucía, Spain), P10-CTS-06068 (Consejería de Innovación, Ciencia y Empresa, Junta de Andalucía, Spain), and Red de Investigación Cardiovascular (RIC) from Redes Temáticas de Investigación Cooperativa (RETIC)
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Male ,Aortic valve ,Pathology ,Gene Identification and Analysis ,Myocardial Infarction ,Chemicals and Drugs::Nucleic Acids, Nucleotides, and Nucleosides::Nucleic Acids::DNA::DNA, Single-Stranded::DNA, Complementary [Medical Subject Headings] ,Gene Expression ,lcsh:Medicine ,Vascular Medicine ,Phenomena and Processes::Genetic Phenomena::Genetic Processes::Gene Expression [Medical Subject Headings] ,Cohort Studies ,Anatomy::Cardiovascular System::Blood Vessels::Arteries::Aorta [Medical Subject Headings] ,Organisms::Eukaryota::Animals::Chordata::Vertebrates::Mammals::Primates::Haplorhini::Catarrhini::Hominidae::Humans [Medical Subject Headings] ,Bicuspid aortic valve ,Reference genes ,Gene expression ,Morphogenesis ,Medicine and Health Sciences ,lcsh:Science ,Aorta ,Anatomy::Cardiovascular System::Heart::Heart Valves::Aortic Valve [Medical Subject Headings] ,Multidisciplinary ,Algoritmos ,Nuclear Proteins ,RNA-Binding Proteins ,Middle Aged ,Congenital Heart Defects ,Neoplasm Proteins ,Humanos ,medicine.anatomical_structure ,Cardiovascular Diseases ,Research Design ,Diseases::Pathological Conditions, Signs and Symptoms::Pathological Conditions, Anatomical::Dilatation, Pathologic [Medical Subject Headings] ,cardiovascular system ,Female ,RNA extraction ,Anatomy ,Algorithms ,Cyclin-Dependent Kinase Inhibitor p27 ,Dilatation, Pathologic ,Research Article ,Phenomena and Processes::Chemical Phenomena::Biochemical Phenomena::Biochemical Processes::Nucleic Acid Denaturation [Medical Subject Headings] ,Dilatación Patológica ,medicine.medical_specialty ,Expresión Génica ,Clinical Research Design ,Health Care::Population Characteristics::Demography [Medical Subject Headings] ,Cardiology ,Aortic Diseases ,Desnaturalización de Ácido Nucleico ,Diseases::Cardiovascular Diseases::Vascular Diseases::Aortic Diseases::Aortic Aneurysm [Medical Subject Headings] ,In Vitro Techniques ,Biology ,Real-Time Polymerase Chain Reaction ,Research and Analysis Methods ,Molecular Genetics ,Diseases::Cardiovascular Diseases::Heart Diseases::Heart Valve Diseases [Medical Subject Headings] ,Anatomy::Tissues [Medical Subject Headings] ,medicine.artery ,Valvular Diseases ,Enfermedades de las Válvulas Cardíacas ,Aneurisma de la Aorta ,Ascending aorta ,Genetics ,medicine ,Humans ,Demografía ,Birth Defects ,ADN Complementario ,Gene ,Aged ,Congenital Hereditary Myopathies ,Clinical Genetics ,Acute Cardiovascular Problems ,lcsh:R ,Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Genetic Techniques::Nucleic Acid Amplification Techniques::Polymerase Chain Reaction::Real-Time Polymerase Chain Reaction [Medical Subject Headings] ,Biology and Life Sciences ,Analytical, Diagnostic and Therapeutic Techniques and Equipment::Investigative Techniques::Genetic Techniques::Nucleic Acid Amplification Techniques::Polymerase Chain Reaction [Medical Subject Headings] ,Human Genetics ,medicine.disease ,Reacción en Cadena en Tiempo Real de la Polimerasa ,Válvula Aórtica ,Cardiovascular Anatomy ,Reacción en Cadena de la Polimerasa ,lcsh:Q ,Information Science::Information Science::Computing Methodologies::Algorithms [Medical Subject Headings] ,Developmental Biology - Abstract
Journal Article; Dilatation of the ascending aorta (AAD) is a prevalent aortopathy that occurs frequently associated with bicuspid aortic valve (BAV), the most common human congenital cardiac malformation. The molecular mechanisms leading to AAD associated with BAV are still poorly understood. The search for differentially expressed genes in diseased tissue by quantitative real-time PCR (qPCR) is an invaluable tool to fill this gap. However, studies dedicated to identify reference genes necessary for normalization of mRNA expression in aortic tissue are scarce. In this report, we evaluate the qPCR expression of six candidate reference genes in tissue from the ascending aorta of 52 patients with a variety of clinical and demographic characteristics, normal and dilated aortas, and different morphologies of the aortic valve (normal aorta and normal valve n = 30; dilated aorta and normal valve n = 10; normal aorta and BAV n = 4; dilated aorta and BAV n = 8). The expression stability of the candidate reference genes was determined with three statistical algorithms, GeNorm, NormFinder and Bestkeeper. The expression analyses showed that the most stable genes for the three algorithms employed were CDKN1β, POLR2A and CASC3, independently of the structure of the aorta and the valve morphology. In conclusion, we propose the use of these three genes as reference genes for mRNA expression analysis in human ascending aorta. However, we suggest searching for specific reference genes when conducting qPCR experiments with new cohort of samples. Yes
- Published
- 2014
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