| Pear fruit cv.’Nanguo’(Pyrus ussuriensis Maxim.) are native to Liaoning province and harvest before the middle of September annually. Nanguo pears are unfavorable for storage at room temperature after harvest. They can just store for about 20 days at room temperature after harvest, and then senesce fast with low product value. Refrigeration is an effective method to delay ripening and reduce decay of Nanguo pears, and it can prolong their storage life for 6 months.1-MCP (1-methylcyclopropene) is an ethylene receptor signal transduction inhibitor and 1-MCP treatment is an effective method for the long-term storage of fruit. Aroma quality is important to Nanguo pears, but refrigeration and 1-MCP treatment can both result in a loss of aroma of Nanguo pears. So to study on the mechanism of the effects of refrigeration and 1-MCP treatment on the aroma of Nanguo pears and explore appropriate controlling methods are of great significance. In this study, Nanguo pears were subjected to refrigeration and 1-MCP treatment and changes in aroma-related esters, levels of fatty acids, the activity and gene expression of alcohol acyltransferase (AAT), lipoxygenase (LOX), and alcohol dehydrogenase (ADH) were investigated. Changes in the activity and gene expression of enzymes involved in ethylene biosynthesis and the expression of genes involved in ethylene signal transduction were also studied. In addition, intermittent warming (IW) and low temperature conditioning (LTC) treatments were applied to study the regulation on the aroma of refrigeration and 1-MCP treated Nanguo pears. The main research results are as follows:From Nanguo pears, cDNAs were isolated from two aroma-related genes, PuAAT1 (GenBank Accession No.KJ775788) and PuLOXl (GenBank Accession No.KJ775789). The cloned cDNA sequence of PuAAT1 contained an open reading frame (ORF) with 1380 bp and encoded a polypeptide of 459 amino acids. The partial length of PuLOXl cDNA encodes a polypeptide of 423 amino acids. PuAATl has four potential.N-glycosylation sites, one protein kinase C phosphorylation sites, seven CK-2 phosphorylation sites and one.N-myristoylation sites. And there exist a phosphorylation site composed of nine serines, three threonines and five tyrosines. Most amino acid sequences of PuAATl are hydrophilic structure and PuAAT1 amino acid sequence has potential membrane-spanning domains, located at 148th and 174th amino acids. PuAAT1 exhibits the features of other plant acyl transferases, including the HXXXD motif, which is the highly conserved active site consensus sequence present in BAHD family members, and another highly conserved motif FGWG. And PuLOX1 exhibits multiple conservative amino acid area. BLAST for homology search revealed PuAATl to share high similarity with pears and apples. PuLOX1 shared high similarity with LOX genes of pears, apricots and peaches.Refrigeration resulted in a significant reduction in the kinds and production of aroma-related esters during refrigeration and the optimal taste period of the shelf life after refrigeration, and after long-term storage it showed a greater decline. The production of ethylene showed the same results. Low levels of PuACSl expression during refrigeration and low levels of activity and expression of 1-aminocyclopropane-l-carboxylate (ACC) synthase (ACS) and ACC oxidase (ACO) resulted in low ethylene production. And the expression of genes involved in ethylene signal transduction (PuETR1 and PuCTRl) was inhibited by refrigeration. During the refrigeration, low temperature had no significant impact on activity of LOX and AAT and expression of PuADH3, but activity of ADH and expression of PuLOX1 and PuAATl declined. During the optimal taste period of shelf life after these storage periods, the loss of aroma-related esters in pear fruit was related to the decreased expression and activity of AAT, LOX, ADH, and polyunsaturated fatty acids levels.1-MCP treatment resulted in a significant reduction in the kinds and production of aroma-related esters of Nanguo pears during refrigeration and the optimal taste period of the shelf life after refrigeration. Though 1-MCP treatment had no significant impact on activity of ACS and ACO during the refrigeration and the activity of ACS during the optimal taste period of the shelf life after refrigeration, it inhibited the PuACSl and PuACO1 expression and then made the production of ethylene declined. In addition, PuETR1 and PuCTR1 expression was inhibited by 1-MCP treatment. The loss of aroma-related esters in 1-MCP-treated pears was related to the decreased polyunsaturated fatty acids levels, activity of ADH and AAT, and expression of PuLOXl, PuADH3 and PuAATl.IW and LTC treatments promoted the production of aroma-related esters of refrigerated and 1-MCP-treated pears during refrigeration and the shelf life after refrigeration. IW-treated and LTC-treated pears produced more ethylene while they were at the same level of maturity with their respective control pears. High levels of activity and expression of ACS and ACO in IW-treated and LTC-treated pears may result in the high level of ethylene production. IW and LTC treatments promoted the expression of PuETRl and PuCTRl in refrigerated pears during refrigeration and the shelf life after refrigeration, but there existed differences in effects of IW and LTC treatments on the two genes’expression of 1-MCP-treated pears. The high levels of linoleic and linolenic acids contents may contribute to the formation of aroma-related esters. During refrigeration and the shelf life after refrigeration, there existed differences in effects of IW and LTC treatments on the levels of activity and expression of LOX, ADH and AAT, but the higher levels of activity and gene expression of them may promote the formation of aroma-related esters. |
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