Abstract:

This study aimed at investigating the transcription and expression of recombined plasmid pcDNA3-gtfB which en- coding multiple glucosyltransferase-B antigenic gene, and the feasibility of the pcDNA3-gtfB used as gene vaccine.Methods:The pcDNA3-gtfB was transfected intomammalian cell COS-1 with liposome. The total RNAof COS-1 cell transfected by pcDNA3-gtfB was extracted and purified. Using the total RNAas template, the transcription of pcDNA3-gtfBwas assayed by reverse transcription polymerase chain reaction (RT-PCR). The expression product of pcDNA3-gtfB was identified with 5% SDS-PAGE, and then as- sayed usingWestern-blotting. The expression productof pcDNA3-gtfBwas also assayed by using LSAB method, and cell transfect- ed by pcDNA3 as the negative control.Results:Identified by agarose gel electrophoresis, the target gene fragment had the same molecular size (316 kb) as itwas predicted, and it indicated that pcDNA-3gtfB was correctly transcribed into mammalian cells. Proved by SDS-PAGE, the molecularweightof the expression product(116~212 kD) was also the same as itwas supposed to be. Itwas also indicated by Western-blotting and LSAB assay that the expression product induced immunizing response.Conclusion: As gene vaccine, itis importance that the recombined plasmid could be correctly transcribed and expressed inmammalian cells. It was suggested by RT-PCR, LSAB and Western-blotting that recombined plasmid pcDNA3-gtfB could be correctly transcribed and expressed in mammalian cells, and the expression product could induce immunizing response, which support its use as gene vac- cine candidates in the development of anticaries vaccines.

Key words: anticaries, Streptococcus mutans, glucosyltransferase, gene vaccine