Restriction Analysis of Recombinant Plasmids

A key step in the construction of recombinant plasmids is the verification of the successful cloning of insert DNA into the vector. A number of commonly used plasmids facilitate phenotypic selection and/or screening methods for rapid identification of insert-containing clones. Additional screening methods are required to confirm the presence and/or orientation of the intended insert. A polymerase chain reaction (PCR) strategy may be sufficient to verify the presence and orientation of the intended insert depending on the availability of vector- and insert-specific primers (see Chapter 19 ). In situations where PCR-based analysis is undesirable or additional analysis is required, clear information about the clone can be derived from restriction analysis.