We have used short oligonucleotides to genetically transform the Lyme disease spirochaete The oligonucleotides are derived from the sequence of an Arg-133 to He mutant (chromosomal) gene that confers resistance to the antibiotic coumermycin A. Oligonucleotides were about 10000-fold less efficient at transformation, on a molar basis, than longer PCR-generated substrates. All of the transformants tested contained the predicted site-directed silent mutation in their genes. Antisense oligonucleotides were more efficient at transformation than either sense or double-stranded oligonucleotides. This is the first demonstration of oligonucleotides used to introduce site-directed mutations directly into the genome of a bacterium.


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