Reference Genes for High-Throughput Quantitative Reverse Transcription-PCR Analysis of Gene Expression in Organs and Tissues of Eucalyptus Grown in Various Environmental Conditions

Autor(es)

Cassan-Wang, Hua ^[1] ; Soler, Marcal ^[1] ; Yu, Hong ^[1] ; Camargo, Eduardo Leal O. ^{[1, 2]} ; Carocha, Victor ^{[1, 3, 4]} ; Ladouce, Nathalie ^[1] ; Savelli, Bruno ^[1] ; Paiva, Jorge A. P. ^{[3, 4]} ; Leple, Jean-Charles ^[5] ; Grima-Pettenati, Jacqueline ^[1]

Afiliação do(s) autor(es)

^[1] Univ Toulouse 3, UPS, CNRS, LRSV, F-31326 Castanet Tolosan - France

^[2] Univ Estadual Campinas, UNICAMP, Inst Biol, Dept Genet Evolucao & Bioagentes, Lab Genom & Expr, BR-13081970 Campinas, SP - Brazil

^[3] Inst Invest Cient & Trop IICT MNE, P-1349007 Lisbon - Portugal

^[4] Univ Nova Lisboa, Inst Tecnol Quim & Biol, Lab Biotecnol Celulas Vegetais, P-2781901 Oeiras - Portugal

^[5] INRA, Ameliorat Genet & Physiol Forestieres AGPF UR0588, F-45075 Orleans - France

• Tipo de documento: Artigo Científico
• Fonte: Plant and Cell Physiology; v. 53, n. 12, p. 2101-2116, DEC 2012.
• Citações Web of Science: 36

Resumo

Interest in the genomics of Eucalyptus has skyrocketed thanks to the recent sequencing of the genome of Eucalyptus grandis and to a growing number of large-scale transcriptomic studies. Quantitative reverse transcription-PCR (RT-PCR) is the method of choice for gene expression analysis and can now also be used as a high-throughput method. The selection of appropriate internal controls is becoming of utmost importance to ensure accurate expression results in Eucalyptus. To this end, we selected 21 candidate reference genes and used high-throughput microfluidic dynamic arrays to assess their expression among a large panel of developmental and environmental conditions with a special focus on wood-forming tissues. We analyzed the expression stability of these genes by using three distinct statistical algorithms (geNorm, NormFinder and delta Ct), and used principal component analysis to compare methods and rankings. We showed that the most stable genes identified depended not only on the panel of biological samples considered but also on the statistical method used. We then developed a comprehensive integration of the rankings generated by the three methods and identified the optimal reference genes for 17 distinct experimental sets covering 13 organs and tissues, as well as various developmental and environmental conditions. The expression patterns of Eucalyptus master genes EgMYB1 and EgMYB2 experimentally validated our selection. Our findings provide an important resource for the selection of appropriate reference genes for accurate and reliable normalization of gene expression data in the organs and tissues of Eucalyptus trees grown in a range of conditions including abiotic stresses. (AU)

• Processo FAPESP: 08/53520-3 - Influencia do nitrogenio na formacao e qualidade da madeira de eucalipto: analise da expressao genica e identificacao de genes visando o melhoramento genetico.
• Beneficiário: Eduardo Leal Oliveira Camargo
• Modalidade de apoio: Bolsas no Brasil - Doutorado