Mutations can also be generated by insertional mutagenesis. The components of this recombination system were elucidated using deletion mutagenesis studies. This process is referred to as insertional mutagenesis or transposon mutagenesis. Then they performed mutagenesis and functional expression, where plasmid constructs were transcribed and expressed. Retroviruses and transposons are the chief instrumental tools in insertional mutagenesis. Absence of cytosolic SOD causes a dramatic increase in mutagenesis and genomic instability. Recently many methods for transposon based random mutagenesis have been reported. Random mutations can be introduced using either error prone PCR, or site saturation mutagenesis. This is advantageous as insertional mutagenesis is avoided. Apoptosis can prevent cells with excess DNA damage from undergoing mutagenesis and progression to cancer. This application has novel implications in site-specific mutagenesis and gene therapy. Recursive ensemble mutagenesis then utilizes information from previous combinatorial cassette mutagenesis. First it involves the "in vivo" chemical mutagenesis of the gene of interest via EMS. It has been applied to site-directed mutagenesis and Sanger sequencing. A number of methods exist for the production of these random sequences, including cassette mutagenesis.