本文關(guān)鍵詞： 哈密瓜 冷害 耐冷性 CBF 低溫預(yù)貯 一氧化氮　出處：《西北農(nóng)林科技大學(xué)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：哈密瓜(Cucumis melo L.)肉質(zhì)松脆,風(fēng)味濃郁,營(yíng)養(yǎng)豐富,深受國(guó)內(nèi)外消費(fèi)者的喜愛,是新疆重要的園藝特產(chǎn)之一。由于其采收季節(jié)溫度高,采后常溫貯藏極易變質(zhì)腐爛。低溫冷藏雖能延緩哈密瓜果實(shí)衰老,但其對(duì)低溫敏感,不適宜的低溫易導(dǎo)致冷害發(fā)生,嚴(yán)重降低了果實(shí)的品質(zhì),影響其商品附加值,限制了低溫冷藏及冷鏈物流技術(shù)在哈密瓜采后保鮮領(lǐng)域的應(yīng)用與推廣。因此,研究哈密瓜果實(shí)采后冷害表現(xiàn)特點(diǎn)及控制方法,已成為哈密瓜產(chǎn)后產(chǎn)業(yè)亟待解決的關(guān)鍵技術(shù)問題。本文以當(dāng)?shù)卮硇云贩N早熟‘西周密25號(hào)’、中熟‘新密3號(hào)’及晚熟‘新密11號(hào)’為試材,研究3±0.5℃貯藏條件下不同品種哈密瓜果實(shí)采后冷害表現(xiàn)特點(diǎn)、耐冷性差異及轉(zhuǎn)錄因子CBF(C-repeatbinding factor)的表達(dá)特性,探討低溫預(yù)貯(low temperature conditioning LTC)和一氧化氮(Nitric oxide NO)對(duì)哈密瓜果實(shí)采后冷害的控制作用及其內(nèi)部機(jī)制,以期為哈密瓜果實(shí)采后耐冷性評(píng)價(jià)體系的建立奠定基礎(chǔ),為生產(chǎn)中冷害的識(shí)別及控制提供理論和技術(shù)依據(jù)。主要研究結(jié)果如下:1.不同品種哈密瓜果實(shí)采后冷害表現(xiàn)特點(diǎn)及采后耐冷性不同�！髦苊�25號(hào)’貯藏21d出現(xiàn)冷害,主要表現(xiàn)為:表皮褐變、凹陷及失水皺縮等癥狀;‘新密3號(hào)’貯藏7d出現(xiàn)冷害,主要表現(xiàn)為:表皮褐變、變色及腐爛等癥狀;‘新密11號(hào)’貯藏42 d出現(xiàn)輕微冷害,主要表現(xiàn)為:表皮點(diǎn)狀褐變等癥狀。不同品種哈密瓜果實(shí)采后冷害主要表現(xiàn)在果皮組織,果肉組織未表現(xiàn)冷害癥狀。冷藏期間,‘新密11號(hào)’果實(shí)冷害指數(shù)和冷害率顯著低于‘西周密25號(hào)’和‘新密3號(hào)’,果實(shí)硬度、可滴定酸(TA)及維生素C含量(Vc)等品質(zhì)指標(biāo)顯著高于后兩者;貯藏49 d后,‘新密11號(hào)’果實(shí)冷害率僅為6.67%,好果率達(dá)到84.44%。表明,‘新密11號(hào)’果實(shí)采后耐冷性強(qiáng)于‘西周密25號(hào)’和‘新密3號(hào)’。2.不同品種哈密瓜果實(shí)采后冷害發(fā)生與果皮組織活性氧代謝、膜脂膜肪酸代謝及內(nèi)源多胺代謝直接相關(guān)。相關(guān)性分析結(jié)果表明,果皮中亞油酸(C18:2)、亞麻酸(C18:3)、腐胺(Put)、精胺(Spm)以及超氧化物歧化酶(SOD)、過氧化物酶(POD)、抗壞血酸過氧化物酶(APX)活性等指標(biāo)與哈密瓜果實(shí)采后耐冷性正相關(guān);乙烯釋放速率、過氧化氫(H_2O_2)、丙二醛(MDA)、超氧陰離子(O_2~(-·))、細(xì)胞膜透性、脂氧合酶(LOX)活性及棕櫚酸(C16:0)含量等指標(biāo)與哈密瓜果實(shí)采后耐冷性負(fù)相關(guān)。因此,建議將以上指標(biāo)作為哈密瓜果實(shí)采后耐冷性評(píng)價(jià)的重要參考指標(biāo),同時(shí)也可將其作為哈密瓜采后果實(shí)冷害發(fā)生的預(yù)測(cè)性指標(biāo)。3.從哈密瓜果實(shí)中分離并克隆獲得轉(zhuǎn)錄因子Cm CBF1(Gen Bank登錄號(hào):KT737742)和Cm CBF3(Gen Bank登錄號(hào):KT737743),分別編碼212個(gè)氨基酸和204個(gè)氨基酸。序列分析表明,Cm CBF1和Cm CBF3包含有一個(gè)高度保守的AP2/EREBP DNA結(jié)合結(jié)構(gòu)域,并含有CBF在AP2/ERF兩側(cè)都含有高度保守的CBF信號(hào)序列及其它特征性結(jié)構(gòu)域。系統(tǒng)進(jìn)化樹分析結(jié)果顯示,Cm CBF1與Md CBF1親緣關(guān)系較近,Cm CBF3與Cs CBF3親緣關(guān)系較近。q RT-PCR分析結(jié)果顯示,不同品種哈密瓜果實(shí)轉(zhuǎn)錄因子Cm CBF1和Cm CBF3均受低溫誘導(dǎo)特異性表達(dá),果皮及果肉中Cm CBF1和Cm CBF3分別在低溫誘導(dǎo)6 h和12 h出現(xiàn)短時(shí)表達(dá)高峰,在貯藏14 d和21 d出現(xiàn)長(zhǎng)時(shí)表達(dá)高峰,Cm CBF3對(duì)低溫反應(yīng)更強(qiáng)烈�！旅�11號(hào)’果皮及果肉中的Cm CBF1和Cm CBF3相對(duì)表達(dá)量顯著高于‘西周密25號(hào)’和‘新密3號(hào)’。相關(guān)性分析結(jié)果表明,轉(zhuǎn)錄因子CBF1和Cm CBF3的表達(dá)與哈密瓜果實(shí)采后耐冷性正相關(guān)。4.研究了不同LTC(10℃預(yù)貯3 d、6 d及9 d轉(zhuǎn)至3℃貯藏)對(duì)‘西周密25號(hào)’果實(shí)采后冷害指數(shù)、冷害率、失重率、好果率及品質(zhì)指標(biāo)的影響。結(jié)果發(fā)現(xiàn),LTC(10℃預(yù)貯3 d后轉(zhuǎn)至3℃貯藏)可顯著抑制冷藏期間哈密瓜果實(shí)呼吸速率和乙烯釋速率,提高果皮中SOD、POD、CAT及APX活性,減少O_2~(-·)生成速率和H_2O_2含量,降低相對(duì)電導(dǎo)率和MDA含量;同時(shí)誘導(dǎo)果皮Cm CBF1及Cm CBF3基因的表達(dá),使果皮中脫落酸(ABA)、生長(zhǎng)素(IAA)及赤霉素(GA)含量保持較高的水平,抑制了多酚氧化酶(PPO)和LOX活性,降低了果皮的水分蒸發(fā),從而減輕‘西周密25號(hào)’果實(shí)冷藏期間表皮凹陷、褐變及皺縮失水等冷害癥狀,使其保持較高的果實(shí)硬度、可溶性固形物、維生素C含量,貯藏49 d后,失重率僅為6.85%,好果率達(dá)40.67%。5.研究了不同NO(0、20、60及100μL/L)處理對(duì)‘新密11號(hào)’果實(shí)采后冷害指數(shù)、冷害率、失重率、好果率及品質(zhì)指標(biāo)的影響。結(jié)果顯示,60μL/L NO處理可緩解‘新密11號(hào)’果實(shí)采后冷害癥狀,并使其冷害發(fā)生推遲7 d,貯藏49 d后,冷害率比對(duì)照組低26.67%。NO處理通過促進(jìn)‘新密11號(hào)’果皮中Put、Spm及亞精胺(Spd)含量的積累,進(jìn)而抑制果實(shí)乙烯的合成,使乙烯峰出現(xiàn)的時(shí)間推遲7 d,從而緩解并推遲果實(shí)采后冷害的發(fā)生;同時(shí)顯著提高冷藏期間果皮中SOD、POD、CAT及APX活性,促進(jìn)游離脯氨酸、可溶性蛋白及可溶性糖等滲透調(diào)節(jié)物質(zhì)的積累,維持較低的O_2~(-·)生成速率和H_2O_2含量,進(jìn)而降低活性氧對(duì)細(xì)胞膜的損傷,維持膜結(jié)構(gòu)穩(wěn)定性及膜系統(tǒng)完整性,減輕哈密瓜果實(shí)在低溫脅迫下的冷害癥狀。另外,NO通過誘導(dǎo)果皮中Cm CBF1及Cm CBF3的表達(dá),提高了哈密瓜果實(shí)采后抗冷性,從而使其保持較好的貨架期品質(zhì)。
[Abstract]:Melon (Cucumis melo L.) flesh crisp, rich flavor, rich nutrition, by domestic and foreign consumers, is one of the important horticultural products in Xinjiang. Because of the high temperature seasons, postharvest storage at room temperature is extremely easy to decay. Low temperature can delay the aging of Hami melon fruit, but it is sensitive to the temperature, not suitable for the low temperature is easy to cause the occurrence of chilling injury, decrease the fruit quality and the effect of the additional value of the goods, limiting the cryogenic cold chain logistics technology and application in the field of fresh Hami melon. Therefore, research on melon postharvest chilling injury characteristics and control methods, has become a key technical problem to be solved after the melon industry. The local representative early-maturing "West careful No. 25 ',' Xinmi 3 'medium and late' Xinmi 11 'as test material, study of 3 - 0.5 DEG C storage under the condition of different varieties The characteristics of symptoms of chilling injury of Postharvest Hami melon, difference in cold tolerance and transcription factor CBF (C-repeatbinding factor) expression, explore the low temperature storage (low temperature conditioning LTC) and nitric oxide (Nitric oxide NO) and the internal mechanism of melon postharvest chilling injury control, in order to lay the foundation for establishment of cold tolerance evaluation system for melon fruit after harvest, to provide theory and technology for identification and control in the production of cold basis. The main research results are as follows: the chilling characteristics and mining 1. different varieties of melon fruit after cold tolerance. "West careful 25 'storage 21d damage, mainly for skin browning, depression shrinkage and other symptoms;' Xinmi 3 'storage 7d damage, the main performance is: Browning and decay, discoloration and other symptoms;' Xinmi 11 'storage 42 d slight damage, the main table Now: skin browning symptoms. Like different fruit varieties of Hami melon chilling injury mainly in peel tissue, pulp tissue showed no chilling injury symptoms. During cold storage, 'Xinmi 11' fruit chilling injury index and chilling injury rate was significantly lower than that of the "west well No. 25 'and' Xinmi 3 ', fruit firmness, titratable acid (TA) and the content of vitamin C (Vc) and other quality indicators were significantly higher than that of the latter two; after 49 D storage,' Xinmi 11 'chilling injury rate was only 6.67%, good fruit rate of 84.44%. showed that the' Xinmi 11 'postharvest resistance to oxygen metabolism of chilling cold and strong in the" West No. 25 well' and 'Xinmi 3'.2. of different varieties of melon fruit and pericarp tissue activity, is directly related to the polyamine metabolism membrane fatty acid metabolism and endogenous. Correlation analysis results showed that the pericarp of linoleic acid (C18:2), linolenic acid (C18:3), putrescine (Put), spermine (Spm) and superoxide Dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX) activity index and melon postharvest cold tolerance positive correlation; ethylene release rate, hydrogen peroxide (H_2O_2), malondialdehyde (MDA), superoxide anion (O_2~ (-)), cell membrane permeability lipoxygenase (LOX) activity and palmitic acid (C16:0) content and the indices such as melon postharvest cold tolerance negative correlation. Therefore, suggestions will be an important reference index for the above indexes as melon postharvest cold tolerance evaluation, but also can be used as a predictive index of.3. occurrence of chilling injury in Hami melon. After isolated and cloned transcription factor Cm CBF1 from melon fruit (Gen Bank accession number: KT737742) and Cm CBF3 (Gen, Bank accession number: KT737743), respectively, encoding 212 and 204 amino acids. Sequence analysis showed that Cm CBF1 and Cm CBF3 contains a highly conserved AP2/EREBP DNA binding domain, and contains a CBF CBF signal contains highly conserved sequence and other characteristic domains on both sides of the AP2/ERF. Phylogenetic analysis showed that Cm CBF1 and Md CBF1 near Cm CBF3 and Cs related, CBF3 is closely related to.Q RT-PCR analysis showed that the different varieties of melon fruit transcription factor Cm CBF1 and Cm CBF3 were affected by the specific expression induced by low temperature, peel and pulp in Cm CBF1 and Cm CBF3 respectively in 6 h and 12 h low temperature induced short-term peak expression, the peak expression in 14 d and 21 d long time storage, Cm CBF3 on the low temperature reaction more intense. 'Xinmi 11' and peel Cm CBF1 and Cm in the flesh of the relative expression of CBF3 was significantly higher than that of the western "careful No. 25" and "Xinmi No. 3". The results of correlation analysis showed that the study of intolerance is positively related to.4. cold expression of transcription factor CBF1 and Cm CBF3 and melon fruit after harvest 鍚孡TC(10鈩冮璐，

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