A method for isolating a polynucleotide of interest that is present in a genome of a first mycobacterium strain or that is expressed by said first mycobacterium strain and that is absent or altered in a genome of a second mycobacterium strain that is different from the first mycobacterium strain or that is not expressed in the second mycobacterium strain, said method comprising:
- a) contacting under hybridizing conditions the genomic DNA of the first mycobacterium strain with the DNA of at least one clone that belongs to a bacterial artificial chromosome (BAC) genomic DNA library of the second mycobacterium strain;
- and
- b) isolating the polynucleotide of interest that fails to form a hybrid with the DNA of the second mycobacterium strain.