DETECTING DNA DAMAGE IN INDIVIDUAL GERM CELLS

A RECENTLY DEVELOPED MICROGEL ALKALINE ELECTROPHORESIS TECHNIQUE PROVIDES AN EXTREMELY SENSITIVE METHOD FOR EVALUATING THE PRESENCE OF DNA DAMAGE (SINGLE-STRAND BREAKS AND/OR ALKALI-LABILE SITES) IN INDIVIDUAL CELLS. BECAUSE EXCISION REPAIR PROCESSES ARE NOT REQUIRED TO DEMONSTRATE THE PRESENCE OF DNA LESIONS AND BECAUSE SAMPLE SIZES CAN BE EXTREMELY SMALL (1 TO <10(5)), THE SINGLE-CELL GEL TECHNIQUESHOULD BE SUPERIOR TO OTHER METHODS CURRENTLY USED TO EVALUATE DNA DAMAGE IN GERM CELLS. FURTHERMORE, BECAUSE DATA ARE COLLECTED AT THE LEVEL OF THE SINGLE CELL, AN ASSESSMENT OF INTERCELLULAR DIFFERENCES IN DAMAGE CAN BE CONDUCTED, MAKING IT MORE INFORMATIVE THAN BIOCHEMICAL TECHNIQUES BASED ON POOLED CELL SAMPLES. THE PURPOSE OF THIS PROJECT IS TO EVALUATE THE SUITABILITY OF THIS TECHNIQUE FOR DETECTING DNA DAMAGE IN THE CELLS OF MALE MICE EXPOSED TO POSSIBLE MUTAGENS. THE FOCUS OF THE PROJECT WILL BE ON THE VARIOUS METHODS USED TO ISOLATE GERM CELLS FROM THE TESTIS AND THE KINETICS OF THE INDUCTION OF DNA DAMAGE INDUCED BY SELECTED MUTAGENIC AGENTS (ACRYLAMIDE,CIS-PLATINUM, CYCLOPHOSPHAMIDE, AND METHYLMETHANESULFONATE).