II.27. Kinetic studies on the induction of the inhibition of repair of radiation damage in barley.
C. Sander, A. Kleinhofs, C, Konzak and R, A, Nilan, Department of Agronomy and Soils, Washington State University, Pullman, Washington 99163, U.S.A.
The respiratory inhibitor sodium azide has been used to probe the nature of repair of radiation induced genetic damage in barley. Soaking non-irradiated seeds in azide solution reduces seedling height, provided the treatment is below the pK (4.7) of azide. Thus, 10-3 M azide (pH 3, 2 hrs,, 20°C) reduces the seedling height at five days to 70-80% of controls. This same treatment produces a relatively high frequency of M2 chlorophyll deficient mutants (15 mutants/100 spikes). The number of these mutants is increased with increasing presoak times until a plateau of 45 mutants/100 spikes is obtained, in treatments involving two or more hours of presoaking. The induction of mutations is unaffected by 02 or N2 bubbling during the treatment. The seedling heights showed no significant changes with varying presoak (up to 4 hrs.).
Azide treatment (10-3M, 2 hrs, 20°C) has a marked effect of the seedlings heights from irradiated seeds (gamma rays, 20 kR). Thus, below its pK, azide reduces the seedling height to 35% of the irradiated control. The reduction in seedling height closely parallels the acid titration curve of sodium azide. At neutral and alkaline pH, azide is ineffective in reducing seedling heights.
Azide does not appear to produce chromosome aberrations per se. However, the chromosome aberration frequencies obtained with azide-treated irradiated seeds show a strong synergistic effect. The total number of aberrations scored at anaphase after a 2 hr, treatment by azide (10-3M) showed a 50% increase over irradiated control.
Repair of radiation damage is apparent when irradiated seeds are allowed to imbibe buffer prior to azide treatment, Thus, the total number of aberrations is reduced if presoaking is extended to one hour and involves air or 02 bubbling. With prolonged presoaks, the reduction in aberrations is no longer apparent. No reduction in aberrations is observed in anaerobic (N2) systems.
Preliminary analysis of single-strand breaks in DNA indicate a correlation between chromosomal repair and rejoining at DNA level, It is expected that further analysis of DNA breaks following azide and radiation treatments will help determine if mutations and chromosome breaks are induced and repaired by the same basic mechanisms.