ABSTRACT
Molecular cloning strategies depend largely upon what is already known about the protein or gene of interest, the size and abundance of the mRNA, and the availability of specific screening tools. Strategies beyond the scope of this brief overview include differential and subtractive cloning methods, functional and ligand screening assays, and polymerase chain reaction techniques for target genes where no sequence information is known. Straightforward polymerase chain reaction strategies are highlighted that can enhance progress in cooperative structural studies. Results of the SVS II study clearly illustrate the complementarity of protein chemistry and molecular biology. Direct analyses of this prototype plant toxin have revealed the mature protein structure and provided a sequence information critical for cloning, whereas DNA studies have demonstrated that the A and B chains are synthesized together as a single polypeptide containing both leader and linker sequences.
