Transformation of Saccharopolyspora spinosa protoplasts with plasmid DNA modified in vitro to avoid host restriction

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Abstract
Summary: Saccharopolyspora spinosa protoplasts were not transformable by several different streptomycete plasmids, and S. spinosa was not a host for plaque formation by the Saccharopolyspora bacteriophages ØSE60, ØSE45, ØSE57, ØSE60, ØSE69 or HP10. Extracts of S. spinosa contained DNA-modifying activities that blocked cleavage of plasmid DNA by Nael and Sail, and partially blocked cleavage by Ncol. Plasmid pOJ434, a derivative of the S. spinosa plasmid pSAS1, was modified in vitro by incubation with an extract of S. spinosa to circumvent restriction. Under optimal conditions, S. spinosa protoplasts were transformed to apramycin resistance by modified pOJ434 at frequencies of about 104 per μg of DNA. Transformants contained pOJ434 primarily integrated in the chromosome.

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