Construction of eag deletion mutant of Bacillus anthracis vaccine strain A16R

Abstract:

[Objective] Construction of eag deletion mutant of Bacillus anthracis vaccine strain A16R. [Methods] To study the function of the gene eag of Bacillus anthracis vaccine strain A16R, according to the sequence of Bacillus anthracis Ames strain, we designed primers and constructed a recombinant plasmid by the spectinomycin resistance cassette, upstream homologous fragment and downstream homologous fragment of eag cloned in tandem in pKSV7. We introduced the recombinant into A16R by electroporation and screened the mutant using the principle of homologous recombination. We checked the mutant using the PCR and proteomics. [Results] We constructed the recombinant plasmid successfully and got the eag deletion mutant. PCR results showed the gene eag was deleted; SDS PAGE showed evident differences between prime strain and mutant strain. Two-dimensional gel electrophoresis results displayed three protein points identified as EA1 by mass chromatographic analysis presented in prime strain had absented from the mutant strain. [Conclusion] We constructed eag deletion mutant of Bacillus anthracis vaccine strain A16R?. This research will be helpful to study the functions of eag gene and the functions of the important genes of Bacillus anthracis.