Functional analysis of pAL5000, a plasmid from Mycobacterium fortuitum: construction of a "mini" mycobacterium-Escherichia coli shuttle vector.
Functional domains of pAL5000 were determined by gene disruption and deletion analysis. Of the five plasmid open reading frames (ORFs), ORF1 to ORF5, and a putative origin of replication previously identified (J. Rauzier, J. Moniz-Pereira, and B. Gicquel-Sanzey, Gene 71:315-321), two of the ORFs (ORF3 and ORF4) were deemed dispensable for plasmid replication. A "mini" mycobacterium-Escherichia coli shuttle plasmid applicable for general recombinant DNA studies in mycobacteria was constructed by using the gene for Kanr (Tn903) as a selective marker. Heterologous expression of the gene for Kanr was confirmed by Western blotting (immunoblotting) analysis.
Ranes
M G
MG
Unité de Génie Microbiologique, Institut Pasteur, Paris, France.
Rauzier
J
J
Lagranderie
M
M
Gheorghiu
M
M
Gicquel
B
B
D002872:Chromosome Deletion; D004261:DNA Replication; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors; D015151:Immunoblotting; D009161:Mycobacterium / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D015183:Restriction Mapping; D013045:Species Specificity
D004251:DNA Transposable Elements; D004269:DNA, Bacterial