1. INHIBITION OF ETHYLENE IN APRICOT FRUIT: ACTION VERSUS BIOSYNTHESIS González-Agüero, M 1,2., Rubio, P. 3, Pizarro, M. 3, Gudenschwager, O 1.Campos-Vargas, R. 1,2 and Defilippi, B.G. 1,2 *. 1 Instituto de Investigaciones Agropecuarias, INIA-La Platina. Casilla 439-3, Santiago 2 The Plant Cell Biotechnology Millennium Nucleus (PCB-MN) 3 Facultad de Ciencias Agronómicas, Universidad de Chile Experimental design Results Conclusions Apricot (Prunus armeniaca L.) fruit is highly susceptible to flesh softening and loss of flavor, particularly during postharvest storage. Most of these changes are under ethylene regulation. During the last two seasons we have studied the effect of 1-methylcyclopropene and aminoethoxyvinylglycine (an ethylene action and an ethylene synthesis inhibitor, respectively) on quality attributes of Patterson apricot cultivar. Both ethylene inhibitors have been effective in reducing ethylene production, fruit softening and color development. On the other hand, soluble solids concentration and titratable acidity have shown an ethylene-independent pattern. In order to understand the effect of both inhibitors in the ethylene biosynthetic pathway, we identified, cloned and characterized the expression pattern of the key genes involved in ethylene synthesis and perception (ACS, ACO, ETR and ERS). The expression profile was characterized by qPCR as ripening progressed at 20 °C after harvest. The significance of the changes measured during apricot ripening is discussed. 1. Evaluation of quality attributes from two seasons trial. Maturity parameters analyzed included: ethylene production rate (A), fruit firmness (B) and color (C). For treatments information see above. 2. Schematic representation of AVG and 1-MCP effect on ethylene biosynthetic pathway (A). For each gene analyzed we obtained the full length sequence by RACE-PCR. (B) shows the experimental procedure for cloning of the etr gene (example) and possible structure of Pa-etr from P. armeniaca. 3. Gene expression analyses performed in season 2 for acs, aco, etr and ers. Expression patterns for the 6 transcripts were characterized by qPCR in 4 fruits for each treatment. Amplification assays were performed three times. Gene expression was normalized considering an external control (Gene dap from Bacillus subtilis), and expressed as a percentage of the highest value of relative abundance. Funded by Fondecyt 1060179 and PBCT PSD03 - Patterson apricots applied with AVG and 1-MCP displayed a lower ethylene production rate than the non-applied fruit. - Despite that both inhibitors have a different mode of action, we did not observe differences at transcripcional level in the genes studied. - Only acs2 would be regulated by ethylene, reducing its expression significantly with 1-MCP and AVG applications. For 1-MCP, the low level of expression of acs2 could be due to the importance of ethylene action in the autocatalytic production system of climacteric fruits. * Different letters represent significant differences at P < 0.05 by LSD test. * Treatments performed in season 1: T1Control T21000 nL L -1 1-MCP (SmartFresh™) T310000 nL L -1 1-MCP (SmartFresh™) T4 100 mg L -1 AVG (Retain® 15% p/p) T51000 mg L -1 AVG (Retain® 15% p/p) * Treatments performed in season 2: CControl 1-MCP10000 nL L -1 1-MCP (SmartFresh™) AVG1000 mg L -1 AVG (Retain® 15% p/p) Apricot cv. Patterson Treatments * Evaluation of quality attributes Genes analyzed: acs, aco, etr and ers Search of ortholog sequences Full length coding sequences (RACE-PCR) Primers design for qPCR Gene expression analyses of acs, aco, etr and ers Real Time PCR (qPCR) RNA extraction, cDNA synthesis Season 1Season 2 Evaluation of quality attributes - Firmness - Ethylene - Color - Etc. 0°C 0°C + 20°C 20 °C Season 1Season 2 A B C Evaluation time (days) (µL C 2 H 4 / k*h) 20 20 + 4 30 30 + 1 kg - f 20 20 + 4 30 30 + 1 Hue angle 20 20 + 4 30 30 + 1 Evaluation time (days) Harvest 2 4 7  1 (µL C 2 H 4 / k*h) Harvest 2 4 7  1 Kg - f Hue angle * Different letters represent significant differences at P < 0.05 by LSD test.Methionine SAM SAM Ethylene (C 2 H 4 ) ACC Transcription factors ACC synthase 1,2 y 3 (ACS) ACC oxidase (ACO) Receptors (ETR, ERS) X AVG X 1-MCP A B Search of ortholog sequences in other vegetal species related Primers design RACE-PCR Search of conserved motifs AJ890091 ERS1 Prunus domestica, complete cds (1,929 bp) EF585295 ERS1 Prunus salicina, complete mRNA (2,470 bp) AF316534 ERS Prunus persica, complete cds (1,935 bp) NH 2 COOH Cys-4 Cys-6 H N G1 F G2 Hys-353 Hydrophobic domain GAF HK domain Evaluation time (days) % of Maximum acs1acs2acs3 acoetrers Harvest 2 4 7  1 Evaluation time (days)