利用GFP和抗性双类型标记监测联合固氮菌在玉米根际的定殖

USE OF GFP AND ANTIBIOTIC RESISTANCE AS SELECTIVE MARKERS TO MONITOR COLONIZATION OF ASSOCIATED NITROGEN-FIXING BACTERIA IN MAIZE RHIZOSPHERE

将来自质粒pKRP10、pKRP11和pKRP12的氯霉素、卡那霉素和四环素抗性基因分别插入质粒GFPmut2中gfp基因下游的PstI位点 ,得到gfp和不同抗性基因共存的重组质粒 ,转化Enterobactergergoviae 5 7- 7野生型菌株和耐铵工程菌E7后 ,得到既有抗生素抗性又在蓝光下呈现亮绿荧光的菌株 .用它们接种玉米后 ,利用这两种选择标记双重筛选重新分离到的细菌确定了接种菌在玉米幼苗根表面的定殖数目 ,同时用荧光显微镜观测了它们在玉米根表和土壤中的分布 .确证了GFP与抗生素抗性双标记定量环境中微生物的可靠性和GFP标记用于原位监测细菌分布的优越性 .图版 1图 1表 2参

The chloramphenicol gene of plasmid pKRP10, the kanamycin gene of plasmid pKRP11 and tetracycline gene of plasmid pKRP12 were cloned into the Pst I site on the downstream of gfp gene of plasmid GFPmut2 respectively, the recombined plasmids had the gfp and different antibiotic resistance genes . These plasmids were transformed into Enterobacter gergoviae 57 7 and genetically engineered strain E7 respectively, the resulting transformants exhibited antibiotic resistance and appeared bright green under blue light. Based on the two kinds of selective markers above, the numbers of these transformants colonizing on maize roots were confirmed. The colonization of gfp labeled bacteria on the maize roots as well as their distribution in the soil was observed by fluorescent microscopy. Plate 1, Fig 1, Tab 2, Ref 15