摘要
对从玉米茎内分离的 18株固氮菌进行了回接分离试验 ,经多碳源无氮培养基、菌落形态、颜色与REP PCR相结合的方法 ,筛选、鉴定出了两株具有较高乙炔还原活性和能在玉米体内定殖的菌株R1和R6 .把携带 gfp标记基因的质粒pKK2 2 3 GFP和nifH gfp的质粒pMGFP2 .1分别转化R1和R6 ,得到携带 gfp ,且菌落带绿色荧光的转化子R1A和R6A ,以及携带nifH gfp的转化子R1B和R6B .在限菌条件下对R1A和R6A在玉米根内的定殖以及R1B和R6B中nifH gfp融合基因的表达进行了研究 ,结果表明了R1A和R6A主要定殖在根的皮层细胞、胞间隙、中柱细胞和导管内 ,有时在活的细胞内也可检测到 .R1B和R6B中的融合基因因受玉米根内营养以及其它与固氮酶基因表达相关因素的限制 ,很难在玉米根内表达 ,只有在添加了 1‰蔗糖后 ,nifH gfp才能在根内表达 .图 2表 2参
This paper describes inoculation of 18 bacterial strains isolated from maize to maize, isolation, screening and identification of those strains based on morphology and color, using multi carbon medium, and REP PCR. The result indicated that two diazotrophic bacteria of R1 and R6 with high acetylene reduction activity were obtained. The plasmid of pKK223 GFP harboring gfp (reporter gene) was transferred into R1 and R6, respectively, and two transformants were obtained with the overexpression of GFP, designated as R1A and R6A; and pMGFP2.1 harboring nif H gfp was transferred into R1 and R6, and two transformants carrying pMGFP2.1 were designated as R1B and R6B, respectively. Also the research of colonizing R1A and R6A in maize root tissues, and inducing expression of nif H gfp in R1B and R6B was conducted gnotobiotically. The result showed that R1A and R6A were colonized in intercellular spacer, xylem vessels, wall of xylem vessels and parenchyma of stele predominantly. It was difficult for R1B and R6B to express nif H gfp in root tissues because of the limited nutrients and factors related to dinitrogenas gene expression. Only if 1‰ sucrose was supplemented, nif H gfp could be expressed. Fig 2, Tab 2, Ref 15
出处
《应用与环境生物学报》
CAS
CSCD
2001年第3期207-212,共6页
Chinese Journal of Applied and Environmental Biology
基金
国家高科技发展计划 863项目资助
关键词
玉米
内生固氮菌
回接分离
绿色荧光蛋白基因
定殖
根内表达
限菌条件
maize
endophytic diazotrophic bacteria
inoculation and isolation
green fluorescent protein gene
colonization
