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园艺学报 ›› 2017, Vol. 44 ›› Issue (11): 2126-2134.doi: 10.16420/j.issn.0513-353x.2017-0198

• 研究论文 • 上一篇    下一篇

衣藻β–胡萝卜素加酮酶基因在南瓜果实中的瞬时表达

周洋洋1,2,黄河勋1,李俊星1,3,王 瑞1,3,罗少波1,3,吴廷全1,3,钟玉娟1,3,*   

  1. (1广东省农业科学院蔬菜研究所,广州 510640;2华中农业大学园艺林学学院,武汉 430070;3广东省蔬菜新技术研究重点实验室,广州 510640)
  • 出版日期:2017-11-25 发布日期:2017-11-25

Transient Expression of β-carotene Ketolase of Chlamydomonas reinhardtii in Pumpkin Fruit

ZHOU Yangyang1,2,HUANG Hexun1,LI Junxing1,3,WANG Rui1,3,LUO Shaobo1,3,WU Tingquan1,3,and ZHONG Yujuan1,3,*   

  1. (1Vegetable Research Institute,Guangdong Academy of Agricultural Sciences,Guangzhou 510640,China;2College of Horiculture & Forestry Sciences,Huazhong Agricultural University,Wuhan 430070,China;3Guangdong Key Laboratory for New Technology Research of Vegetables,Guangzhou 510640,China)
  • Online:2017-11-25 Published:2017-11-25

摘要:

为了研究β–胡萝卜素加酮酶基因在南瓜中合成酮基类胡萝卜素的功能,采用农杆菌介导瞬
时表达技术,将含有衣藻β–胡萝卜素加酮酶基因CrBKT 的pBI121-CMTPCRBKT 载体和pBI121 对照载
体的农杆菌菌液注入授粉后2、5、10、15 和25 d 的南瓜果实中,3 d 后观察发现,授粉后2 和5 d 的果
实转化后果肉呈微红,其中授粉后5 d 的颜色较深,而10 d 以上的无红色色素积累。经高效液相色谱
(HPLC)分析色素成分,发现积累的红色色素为酮基类胡萝卜素角黄素和虾青素。与对照相比,注入
CrBKT 基因的授粉后2 和5 d 的幼果中的类胡萝卜素含量显著增加,5 d 的幼果约增加了1/3,且含有106.31
μg · g-1 酮类胡萝卜素,其中角黄素占80.65%,虾青素占19.35%,而授粉后10 d 以上的果实类胡萝卜素含
量没有明显变化,也没有酮类胡萝卜素的积累。PCR 扩增表明转化组织中表达了该基因。结果表明,CrBKT
基因只能在授粉后5 d 以内的幼果中表达,并将幼果中的类胡萝卜素转化为酮基类胡萝卜素;随着果实成
熟,菌液在果肉组织中无法渗透而阻碍基因表达。

关键词: 南瓜;衣藻;&beta, &ndash, 胡萝卜素加酮酶;基因;瞬时表达

Abstract:

In this study,transient expression system was used to analyze the function of
β-ketocarotase for biosynthesis of ketocarotenoid in pumpkin fruit. The Agrobacteria that contained
pBI121-CMTPCRBKT vector with CrBKT gene and the control pBI121 vector were individually injected
into the pumpkin fruits at 2,5,10,15 and 25 days after pollination,respectively. It was found that the fruit
pulp was light reddish in 2- and 5- day fruits with darker color in pulp of 5-day fruit while there was no red pigment accumulated in more than 10-day fruit. High performance liquid chromatography(HPLC)
analysis of pigment composition revealed that the accumulated red pigments were canthaxanthin and
astaxanthin. Compared with the control,the carotenoid content in the 2- and 5-day fruits injected with the
CrBKT gene were significantly increased,which increased by about 1/3 in 5-day young fruit and
ketocarotenoid content was 106.31 μg · g-1,of which canthaxanthin accounted for 80.65%,astaxanthin was
19.35%,while the carotenoid content of the fruits in more than 10-day had no obvious change and there
was no accumulation of ketocarotenoid. PCR amplification indicated that CrBKT was expressed in the
transformed tissue. The result showed that the CrBKT gene could be only expressed in less than 5-day fruit
and it could convert carotenoid in young pumpkin into ketocarotenoid. However,the expression of
exogenous gene was discouraged due to the bacteria could not osmotic into the flesh tissue with the fruit
ripening.

Key words: pumpkin; Chlamydomonas reinhardtii;&beta, -carotene ketolase;gene;transient expression

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