JIN Hualin,JIANG Mingliang,LI Xu,et al.Pectin Decomposition Enzyme Gene Cloning and Expression Analysis in Actinidia arguta Fruit[J].Northern Horticulture,2020,44(15):21-26.[doi:10.11937/bfyy.20194625]
软枣猕猴桃果胶分解酶基因克隆及表达量分析
- Title:
- Pectin Decomposition Enzyme Gene Cloning and Expression Analysis in Actinidia arguta Fruit
- Keywords:
- Actinidia arguta; PG; β-gal; PME; gene cloning
- 文献标志码:
- A
- 摘要:
- 以不同软化阶段的软枣猕猴桃果实为试材，采用分光光度法测定了多聚半乳糖醛酸酶（Polygalacturonase，PG）、β-半乳糖苷酶（β-galactosidase，β-gal）和果胶甲酯酶（Pectin methylesterase，PME）活性，采用RT-PCR法对其控制基因克隆，并对不同成熟阶段的表达进行了分析，以期探讨软枣猕猴桃果实软化机理。结果表明：软枣猕猴桃果实软化中PME和β-gal在软化初期出现活性高峰，PG在软化期形成活性高峰，而且成功克隆得到了软枣猕猴桃PG和β-gal的基因序列。NCBI Blast结果表明与中华猕猴桃PG基因和β-gal基因的一致性分别大于95%和90%。PG基因的半定量分析表明PG基因的转录表达与PG活性协同一致，说明在软枣猕猴桃果实中PG基因的表达是转录水平上调控的；而β-gal基因的半定量分析表明β-gal活性波峰结果一致，但是与果实软化过程中β-gal保持较高的酶活性结果不一致，说明β-gal基因的表达不仅受到转录水平的调控，还可能受到翻译水平上的调控。
- Abstract:
- In order to explore the mechanism of Actinidia arguta fruit softening,Actinidia arguta fruit in different softening stages was used as experimental materials.Polygalacturonase activity (PG),β-galactosidase activity (β-gal) and pectin methylesterase activity (PME) were measured by spectrophotometry.The control gene was cloned by RT-PCR,and the expression at different maturity stages was analyzed.It was found that the activity peaks of PME and β-gal during softening of kiwifruit during the softening period,and the peak activity of PG during the softening period.The gene sequences of PG and β-gal from kiwifruit were successfully cloned.NCBI Blast results showed that the identity with Actinidia chinensis Planch.PG gene and β-gal gene were greater than 95% and 90%,respectively.The semi-quantitative analysis of the PG gene showed that the transcriptional expression of the PG gene was synergistic with the PG activity,indicating that the expression of the PG enzyme gene was regulated at the transcriptional level in A.arguta fruits.The results of β-gal activity peaks are consistent,but they are not consistent with the results of β-gal maintaining a higher enzyme activity during fruit softening,which indicates that the expression of β-gal gene is not only regulated by the transcription level,but also may be regulated by the translation level.
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备注/Memo
第一作者简介：金花林（1992-），女，硕士研究生，研究方向为果树栽培生理。E-mail：KathyKim@163.com.责任作者：朴一龙（1962-），男，博士，副教授，硕士生导师，现主要从事果树栽培生理和果实采后生理等研究工作。E-mail：piaoly@ybu.edu.cn.基金项目：国家自然科学基金资助项目（31060254）。收稿日期：2019-12-08