Detection of Psoralen Cross-Link Sites in DNA Modified by Psoralen-Conjugated Oligodeoxyribonucleoside Methylphosphonates

Oligodeoxyribonucleoside methylphosphonates conjugated with derivatives of psoralen cross-link with complementary singlestranded RNA and DNA and with duplex DNA targets when irradiated with long wavelength (365 nm) ultraviolet light. The position of crosslinking between pyranone-side adducts of psoralen-conjugated oligonucleoside methylphosphonates and DNA can be easily detected by treating the photoadduct with 1M aqueous piperidine at 90°C for 30 min, followed by analysis on a polyacrylamide gel run under denaturing conditions. This treatment results in hydrolysis of the methylphosphonate linkages and cleavage of the phosphodiester backbone at the cross-link site. Multiple crosslinking sites were detected with a singlestranded DNA target which contains four contiguous T residues. This may result from looping out of one or two of the T residues.