Development of a Site-Directed Integration Plasmid for Heterologous Gene Expression in Mycoplasma gallisepticum
نویسندگان
چکیده
Deciphering the molecular basis of the interactions between the parasite Mycoplasma gallisepticum and its avian hosts suffers from the lack of genetic tools available for the pathogen. In the absence of well established methods for targeted disruption of relevant M. gallisepticum genes, we started to develop suicide vectors and equipped them with a short fragment of M. gallisepticum origin or replication (oriC MG). We failed to create a disruption vector, although by adding a further short fragment of the M. gallisepticum tufB upstream region we created a "Trojan horse" plasmid. This is fully integrated into the genomic DNA of M. gallisepticum, always at the same site, oriC MG, and is able to carry and express any gene of interest in the genetic background of M. gallisepticum. Successful expression of a heterologous gene was shown with the lacZ gene of E. coli. When used for gene complementation or expression of hybrid genes in M. gallisepticum, a site-specific combined integration/expression vector constitutes an improvement on randomly integrating transposons, which might have unexpected effects on the expression of chromosomal genes.
منابع مشابه
Development of a replicable oriC plasmid for Mycoplasma gallisepticum and Mycoplasma imitans, and gene disruption through homologous recombination in M. gallisepticum.
The genome of Mycoplasma gallisepticum strain R(low) has been sequenced completely, but subsequent genetic studies have been limited by the lack of a replicable vector system. In this study, replicable plasmids were constructed for M. gallisepticum and Mycoplasma imitans using the oriC region upstream from the soj gene. The oriC plasmids of M. gallisepticum (pGTLori) and M. imitans (pMIori) rep...
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عنوان ژورنال:
دوره 8 شماره
صفحات -
تاریخ انتشار 2013