Enhancement of micronuclei frequency in the Tradescantia/micronuclei test using a long recovery time

The Tradescantia/micronuclei test (TRAD/MCN) is a well-validated test for monitoring environmental genotoxicants. These pollutants induce at the early meiotic stage of pollen mother cells chromosome fragments which become micronuclei at the tetrad stage. The standard test protocol requires some hours of exposure of the inflorescences and a recovery time of about 24 hours to reach the early tetrad stage. Since the recovery period represents a critical step of the TRAD/MCN, experiments were performed to establish its length in plants of clone #4430 of the hybrid T. hirsutiflora x T. subacaulis which is widely used in environmental monitoring. The aim of the present research was to ascertain the exact duration of recovery time in order to improve the sensitivity of the TRAD/MCN test. First, studies were performed to select the flowers at the beginning of the meiosis, and then anthers were sampled and studied for a period of 48-86 hours. The complete meiosis in the plants examined required about 80 hours. Second, exposure to genotoxic substances followed by different recovery times was carried out to demonstrate that effectiveness of the TRAD/MCN test is closely related to the duration of the recovery time. The test was carried out by exposing inflorescences to known mutagens (sodium azide and maleic hydrazide) for six hours followed by different recovery times (24-72 hours). The results showed that the frequency of micronuclei in the pollen mother cells increased with the length of the recovery time.