added the solution was transferred to two 5 8 in u00d7 3 in centrifuge tubes and spun in a beckman ti70 rotor for 42 hours at 44k rpm 17 u00b0 c to collect the plasmids the tubes were opened illuminated with ultraviolet light and the lower of the two fluorescent bands was collected by syringe the lower band from each tube was combined and the ethidium bromide was removed by extracting four times with an equal volume of water saturated ice cold n butanol the extracted solution was dialyzed against 4 changes of dna buffer then the nucleic acid was precipitated by the addition of 2 vols of isopropanol and sufficient 5m nacl to reach a final concentration of 0 4m the solution was stored overnight at 20 u00b0 c then centrifuged for 15 min at 15k rpm 0 u00b0 c the supernatant was discarded the pellet was air dried for 15 min then dissolved in 500 ul of 10 mm tris ph 7 5 1 mm edta and stored at 20 u00b0 c the plasmid dna concentration was found to be approximately 100 ug ml 6 digestion of the plasmid library 6 ug 60 ul of the plasmid library was diluted into 450 ul of restriction endonuclease digestion buffer section 2 60 units 7 5 ul of haeiii restriction endonuclease were added and the tube was incubated at 37 u00b0.