Oligonucleotide chip assay for quantification of gamma ray-induced single strand breaks.
An oligonucleotide chip assay was designed for direct quantification of single strand breaks (SSBs) induced by gamma-ray irradiation. The oligonucleotides used were 20-mers, which were short enough to produce only a single strand break within a single oligonucleotide. The two ends of the oligonucleotides were labeled with fluorescein and biotin, respectively. The biotinylated ends of the oligonucleotides were immobilized on a silicon wafer chip treated with (3-aminopropyl)triethoxysilane (APTES), glutaraldehyde, and avidin. The DNA fragments cleaved by gamma-ray irradiation were detected by a laser-induced fluorescence (LIF) detection system. The gamma-ray-induced SSBs were quantified using a calibration curve (fluorescence intensity versus gamma-ray dose) without the need for complicated mathematical calculation based on gel-based separation. The experimentally determined gamma-ray-induced SSBs yield was almost equal to the theoretical value derived from gel electrophoresis of plasmid DNAs and DNA surface coverage.
Ki
Hyeon A
HA
Research Institute of Pharmaceutical Sciences and College of Pharmacy, Seoul National University, Seoul, South Korea.
Kim
Min Jung
MJ
Pal
Sukdeb
S
Song
Joon Myong
JM
D001710:Biotin / Q000378:metabolism; D019921:Biotinylation; D002138:Calibration; D004249:DNA Damage / Q000528:radiation effects*; D004277:DNA, Single-Stranded / Q000032:analysis* / Q000235:genetics / Q000528:radiation effects*; D004307:Dose-Response Relationship, Radiation; D019793:Fluorescein / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D005720:Gamma Rays; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D009841:Oligonucleotides / Q000235:genetics; D012825:Silicon / Q000737:chemistry; D013499:Surface Properties; D013997:Time Factors
D004277:DNA, Single-Stranded; D005456:Fluorescent Dyes; D009841:Oligonucleotides; D001710:Biotin; D019793:Fluorescein; D012825:Silicon