| The risk of a peel disorder,superficial scald,is a major factor influencing ‘Anjou' pear's postharvest and commercial management.Superficial scald is a physiological disorder characterized as brown or black patches on fruit skin that appears during or after storage on certain pear and apple cultivars.It is accept that conjugated trienols(CTols),the oxidation products of ?-farnesene in fruit peel,are toxic to fruit peel and lead to scald development.Superficial scald susceptibility is impacted by a number of factors including temperature during fruit grown,fruit maturity,storage temperature,storage atmosphere,and ethylene biosynthesis or activity.In this study,we pre-found that the superficial scald never appears on the blushed peel of ‘Anjou' fruit during 8 months of storage,which indicated that the development of superficial scald is affect by pre-harvest light expose.Understanding the mechanism and factors affecting scald development are necessary for predicting scald or developing or postharvest procedures to control scald.In this study,influence of preharvest sunlight exposure on ethylene synthesis,respiration rate,fruit firmness,?-farnesene,CTols,and superficial scald development in green and red‘Anjou' pear among separated blushed and shaded peels of unbagged fruit as well as the whole peel of bagged fruit was tested.There were no significant differences in ethylene production rate,respiration rate,fruit ripening capacity between bagged and unbagged fruit,except significant reduce the SSC in bagged fruit.On the other hand,bagging treatment induced superficial scald development via improve ?-farnesene and CTols content.?-farnesene and CTols only accumulate during cold storage period,and degradation in ripening at room temperature,which indicated that the development of superficial scald is affect by the content of ?-farnesene and CTols during cold storage.There were no significant difference in ?-farnesene concentration between the bulshed and the shaded peels of unbagged fruit,while the content of CTols in the blushed peel of unbagged fruit was lower than that shaded peel throughout the experiment period.Many biochemistry data indicated that the blushed peel has much more antioxidant compounds(anthocyanin,polyphone,flavonols,and flavonoids),which inhibited the oxidant of?-farnesene to CTols by the high value of total antioxidant.On the other hand,the high activity of SOD?MDAR?APX?DHAR?GR in the blushed peel of unbagged fruit,preventedthe lipid peroxidation by reactive oxygen species(ROS)including superoxide anion radical(O2-),perhydroxyl radicals(-OH),hydroxyl radical(H2O2),and therefore reduce scald of fruit.In this study,?-farnesene synthesis gene(AFS)was cloned from green and red ‘Anjou'fruit,which was named PcAFS1.The full length cDNA of PcAFS1 is 1952 bp and encodes a protein of 577 amino acids,and the homology is 99.3%.A phylogenetic tree containing PcAFS1 and ?-farnesene or ?-farnesene synthesis genes were constructed with a neighbor-joining method.The result showed that PcAFS1 is highly to apple ?-farnesene synthesis gene.In addition,qRT-PCR analysis showed that PcAFS1 constitutively expressed in all tested organs,with the highest transcript level in ripened pear peel.Meanwhile and its expression is induced by ethylene condition,and positively associated with the accumulation of ?-farnesene synthesis during cold storage.Furthermore,the transcript level of PcAFS1 is inhibited by 1-MCP treatment.The primers were colned in genome of red and green ‘Anjou',the full length primer of PcAFS1 is 2067 bp in both cultives,and the homology is 99.6%,there is only 9 bp differnence.Cis-elements on PcAFS1 primer were analysised by PLANTCARE(http://bioinformatics.psb.ugent.be/webtools/plantcare/html/)online,and HSE(cis-acting element involved in heat stress responsivenss)and MBS(MYB biding site)were found,which indicated that PcAFS1 gene maybe response by high temperature treatment and regulated by MYB transcript factor.To determine the function of PcAFS1,expression constructs containing pBI121-PcAFS1 driven by cauliflower mosaic virus 35 S promoter are genetically transformed into Arabidopsis.PcAFS1 overexpression in Arabidopsis increases the accumulation of ?-farnesene.After treated with ethylene(0.2 ?l l-1),transgenic plantlets produce increased ?-farnesene,and indus leaves bent down.Anthocyanins in light and bagged differences in dealing with the largest,which probably played a key role in the process of superficial scald development.Inorder to uncover the relationship between anthocyanin and superficial scald development,a MYB transcript factor was coloned in red ‘Anjou' fruit,which was named PcMYB1.This gene is only transcript in red ‘Anjou' fruit,not green ‘Anjou'.The primer sequences of PcMYB1 were coloned in both cultivars,and the homology is 99.3%,only 12 bp difference.While the genome sequence show 244-bp lost in the first intron of red ‘Anjou',the allelic sequence lost in the genome of PcMYB1 maybe sufficient to account for the increase in PcMYB1 transcript levels and subsequent ectopic accumulation of anthocyains throughout the fruit.In the first few months of storage,the content of ?-farnesene was higher in red ‘Anjou'than green ‘Anjou'.The transcription level of PcAFS1 was wakeup in 1-MCP treated red‘Anjou' fruit,but completed inhibited in green ‘Anjou'.To further characterize the functionof PcMYB1 regulated PcAFS1 gene expression,35S::PcMYB1,35S::PcbHLH3,35S::PcWD40,and PPcAFS1::LUC were transferred into tobacco leaves.Transient assays demonstrated that PcMYB1 promoted transcript of PcAFS1,while the activity was not increase by co-transcript by bHLH or WD40 transcription factors.This indicated that the mechanism of MYB regulates anthocyanin and ?-farnesene was difference.Taken together,those results indicate that sun-expose increased the accumulation of antioxidant compounds,such as anthocyanin,polyphone,flavonols,and flavonoids,up-regulated the activity of antioxidant enzyme,such as SOD?MDAR?APX?DHAR?GR,which led a natural resistant to superficial scald by the higher antioxidant activity.On the other hand,PcMYB1 overexpression in red ‘Anjou' increased accumulation of ?-farnesene by up-regulated the transcript level of PcAFS1.However,red ‘Anjou' showed less superficial scald index because the higher content of antioxidant such as anthocyanin. |
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