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 single-stranded RNA and DNA and with duplex DNA targets when irradiated with long wavelength (365 nm) ultraviolet light. The position of cross-linking between pyranone-side adducts of psoralen-conjugated oligonucleoside methylphosphonates and DNA can be easily detected by treating the photoadduct with 1 M aqueous piperidine at 90 degrees 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 cross-linking sites were detected with a single-stranded DNA target which contains four contiguous T residues. This may result from looping out of one or two of the T residues.
Kean
J M
JM
Department of Biochemistry, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205.
Miller
P S
PS
D001483:Base Sequence; D001665:Binding Sites; D003432:Cross-Linking Reagents; D004247:DNA / Q000737:chemistry*; D005363:Ficusin / Q000737:chemistry*; D005561:Formates / Q000494:pharmacology; D008969:Molecular Sequence Data; D009705:Nucleosides / Q000737:chemistry*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D009943:Organophosphorus Compounds / Q000737:chemistry*; D010880:Piperidines / Q000494:pharmacology; D011196:Potassium Permanganate / Q000494:pharmacology
D003432:Cross-Linking Reagents; D005561:Formates; D009705:Nucleosides; D009838:Oligodeoxyribonucleotides; D009943:Organophosphorus Compounds; D010880:Piperidines; D011196:Potassium Permanganate; C030544:formic acid; C032627:methylphosphonic acid; C032727:piperidine; D004247:DNA; D005363:Ficusin