Construction and evaluation of a high effective GFP reporter plasmid for Streptococcus pneumoniae / 重庆医科大学学报

ABSTRACT

Objective: To construct a high effective GFP reporter plasmid for screening Streptoccus pneumoniae virulent genes by differential fluorescence induction. Methods: The SD-ENH-GFP region of plasmid pGreenTIR was cloned into a suicide plasmid pEVP3 which contains a cat gene encoding resistance protein to chloramphenicol,and a report plasmid pEVP3-SDGFP was constructed.To evaluate the function of this plasmid,a 500bp fragment of the pneumolysin gene (ply) of TIGR4 was coloned in the upstream of gfp and then was transformed into Streptococcus pneumoniae TIGR4. Results: The plasmid pEVP3-SDGFP could report the expression of ply both in vivo and in vitro,and was more effective than plasmid pEGFP-1 without SD and ENH sequence before gfp gene. Conclusion: Plasmid pEVP3-SDGFP can be used to construct the promoter-trap library which is needed in DFI.