Restriction generated oligonucleotides utilizing the two base recognition endonuclease CwJI

The conversion of an anonymous DNA sample into numerous oligonucleotides is enzymatlcally feasible using an unusual restriction endonuclease, CViJI. Depending on reaction conditions, CWJI is capable of digesting DNA at a two or three base recognition sequence. CviJI normally cleaves RGCY sites between the G and C to leave blunt ends. Under 'relaxed' conditions CWJI* cleaves RGCY, and RGCR/YGCY, but not YGCR sites. In theory, CviJI^* restriction of pUC19 (2686 bp) should produce 157 fragments, 75% of which are smaller than 20 bp. Instead, 96% of the CWJI* fragments were 18-56 bp long and none of the fragments were smaller than 18 bp. Thermal denaturatlon of these fragments generates sequence specific oligonucleotides homologous for the cognate template. The enzymatic conversion of anonymous DNA into sequence specific ollgomers has implications for several conventional and novel molecular biology procedures.