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作物学报 ›› 2014, Vol. 40 ›› Issue (06): 1132-1139.doi: 10.3724/SP.J.1006.2014.01132

• 研究简报 • 上一篇    

绿色荧光蛋白基因标记的固氮菌DX120E在甘蔗植株内的定殖

魏春燕1,邢永秀1,2,*,莫遥1,林丽2,杨丽涛1,2,3,胡春锦3,李杨瑞1,2,3,*   

  1. 1 广西大学农学院 / 亚热带农业生物资源保护与利用国家重点实验室, 广西南宁 530004; 2广西农业科学院 / 中国农业科学院甘蔗研究中心 / 农业部广西甘蔗生物技术与遗传改良重点实验室 / 广西甘蔗遗传改良生物技术重点开放实验室, 广西南宁 530007; 3 广西农业科学院微生物研究所, 广西南宁 530007
  • 收稿日期:2013-10-23 修回日期:2014-03-04 出版日期:2014-06-12 网络出版日期:2014-03-24
  • 通讯作者: 邢永秀, E-mail: document126@126.com; 李杨瑞, E-mail: liyr@gxaas.net
  • 基金资助:

    本研究由国家高技术研究发展计划(863计划)项目(2013AA102604), 国家自然科学基金项目(31171504, 31101122),广西八桂学者、特聘专家专项经费,广西自然科学基金创新团队项目(2011GXNSFF018002), 广西科学研究与技术开发计划项目(桂科产1123008-1, 桂科攻1222009),广西农科院博士后研究专项(86420), 广西农科院团队项目(桂农科2011YT01)和广西自然科学基金(2013GXNSFAA019082)资助。

Colonization of Nitrogen Fixing Bacterial Strain Klebsiella sp. DX120E Labeled with Green Fluorescent Protein (GFP) Gene within Sugarcane Plants

WEI Chun-Yan1,XING Yong-Xiu1,2,*,MO Yao1,LIN Li2,YANG Li-Tao1,2,3,HU Chun-Jin3,LI Yang-Rui1,2,3,*   

  1. 1 State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources / Agricultural College of Guangxi University, Nanning 530004, China; 2 Guangxi Academy of Agricultural Sciences / Sugarcane Research Center, Chinese Academy of Agricultural Sciences / Key Laboratory of Sugarcane Biotechnology and Genetic Improvement (Guangxi), Ministry of Agriculture / Guangxi Key laboratory of Sugarcane Genetic Improvement, Nanning 530007, China; 3 Microbiology Research Institute, Guangxi Academy of Agricultural Sciences, Nanning 530007, China
  • Received:2013-10-23 Revised:2014-03-04 Published:2014-06-12 Published online:2014-03-24

摘要:

为了探索甘蔗固氮菌DX120E菌株侵入根的方式以及菌株在甘蔗植株内的定殖规律, 确定人工接种固氮菌适宜的侵染浓度和定殖特点, 采用绿色荧光蛋白(GFP)标记的固氮菌DX120E接种甘蔗组培苗, 以荧光显微镜观察其侵染和定殖情况, 以抗生素抗性标记法和平板分离计数法测定DX120E菌株的种群动态。结果表明, 菌株DX120E在2个甘蔗品种的根、叶鞘和叶内均能定殖, 定殖菌量依次均为根>叶鞘>叶; 不同接种浓度下, 最大定殖数量无显著差异, 102 CFU mL–1的接种量足够侵入甘蔗并积累定殖DX120E从甘蔗根表面的裂隙、主根和侧根发生处及根的断裂处均可侵入, 主要在根表面细胞间隙和细胞内大量定殖, 同时也可迁移到叶片的叶肉细胞和维管束细胞中定殖。

关键词: 固氮菌(Klebsiella sp.), 甘蔗接种浓度, 细菌数量, 定殖

Abstract:

To explore the invasion patterns of N2-fixing bacteria strain Klebsiella sp. DX120E and its colonization in sugarcane, and to determine the optimum concentrations of inoculum, we inoculated tissue culture seedlings of two sugarcane varieties (GT21 and B8) with different concentrations of Klebsiella sp. bacteria strain DX120E labeled by green fluorescent protein (GFP) gene, and with ddH2O as CK. The results showed that the strain DX120E propagated in root, leaf sheath and leaf of sugarcane seedlings, and bacteria quantities in different parts were root > leaf sheath > leaf. There were no differences in the maximum propagation quantity under various inoculating concentrations, with the optimum concentration of 102 CFU mL–1.Fluorescence microscope observation indicated that the bacteria could invade root through the surface cracks, generating sites of main root and lateral root, and root fracture, and intensively propagate in intercellular spaces and cells of the root, also transfer to mesophyll cells and vascular bundle sheath cells in leaf.

Key words: Nitrogen fixing bacteria, Klebsiella sp., Sugarcane, Inoculation levels, Bacteria quantites, Colonization

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