A total of 45 participants from 13 different countries. Among the participants three were professors from Universities and 40

graduate students most were from Eastern European countries.

International Program on Plant Bioassays /under the auspices of the United Nations Environment Program (IPPB/UNEP) conducted the Hands-on Workshop at the Institute for Cancer Research, Medical University of Vienna, Vienna, AustriaJuly 2006.

Prof. Siegfried Knasmueller, Director of the Institute for Cancer Research, and Prof. Te-H Ma, from Western Illinois University Macomb, IL, USA . And Dr. Miroslav Misik of the Institute for Cancer Research served as instructors of the workshop

St. Stevens, Cathedral, land mark of Vienna

Participants of the Hands-on workshop on plant bioassays conducted at the Institute for Cancer Research, Medical University of Vienna, Vienna, Austria

Front entrance of the Institute for Cancer Research, MEDUNIWIEN, Austria Prof. Sigi Knasmueller, Director (Left) and Research Associates

Dr. T. Ma (left) and Dr. Sigi Knasmueller (right) , Directors of the Workshop giving an Introduction of the International Program on Plant Bioassays and instruction of the laboratory exercise.

This workshop was conducted at the Institute for Cancer Research, The Medical University of Vienna, Vienna, Austria, In collaboration with Western Illinois University under the auspices of the NATO/EST Program

One half of the laboratory exercise instruction class

The other half of the participants in the laboratory exercise

Prof. Te Ma, Instructor, demonstrates micronuclei in a microscope

Tradescantia Clone 4430 (sterile hybrid of T. hirusiflora and T. subcaulis, Blue colored flowers and stamen hairs is the result of dominant blue under the heterozygous (P/p) blue and pink alleles .

About 20 –25 individual plants in clone (not solitary) may be grown in a 6 inch pot with regular potting soil. Young flower buds are the targets for exposure to gas and radiation, and to liquid agents via vascular system of plant cuttings (not whole plant with roots). To promote flower bud growth, 16 h/day light is necessary for year round supply of proper experimental material.

Photomicrograph of micronuclei induced by X-rays from the Trad-MCN assay

MNMN

XMN

Pink mutated cells in the stamen hair from the Trad-SHM assay

Micronuclei (arrow) in Meristem (M) and in F1 (F1) regions of root tips from treated roots

During the break of the laboratory exercise, participants taking pictures in the University campus.

Professor of Nuclear Physics lecturing on principle of ionizing Radiation and radiation induced genetic damage -

Radiation detection - Dosimetry of radiation intensity

At the end of the week long workshop, participants receive a certificate given by Dr. Miroslave Misik, Instructor, while Drs. Ma and Knasmuelller greet the recipient.

Vienna - Hands-on Workshop on Plant Genotoxicity Assays European Environmental Mutagen Society Meeting June 26: Preparation of plant materials – Tradescantia cuttings, Allium root germination. Radioactive source (Sand) calibration. Audio/Video equipments for evening lectures or demonstrations Treatment of Tradescantia cuttings. with chemical agents, Gaseous, liquid and radiation treat for Trad-MCN assay. Trad cuttings to be treated a week ahead for Trad-SHM assay (S02) June 27: Preparation of plant materials, Laboratory equipment, Dissecting microscopes, oil immersion light microscopes, and slides, coverglasses dissecting needles stains, reagents (Ethanol, Acetic acid, Carmine stains, hydrochloric acid, Vegetable oil),Score sheets, lens paper, bloting paper. Registration, living accommodation and meal arrangement.June 28: 8:00 – 8:30 Opening ceremony Power point presentation of three Trad-MCN, SHM, Allium MCN bioassays 8:30- 10:30 Slide preparation - Trad-MCN, Trad-SHM, Allium – root tip Demonstration , Technical exercise, observation of Micronuclei, pink mutations in stamen hairs, Hair counting, average number of hairs per flower, scoring of mutation frequencies. 10:30 – 10:45 Coffee break and QA session 10:45 – 12:00 Trad-MCN slide preparation. Trad-SHM assay, 1st day Mutation scoring 12:00 – 13:00 Lunch 13:00 – 15:00 Slide prep. meiotic stages in Trad. inflorescences (inflos) 15:00 – 15:30 Coffee break, QA session, Discussion 15:30 – 17:00 Identify meiotic stages in relation to the flower bud sizes. 17:00 – 19:00 Dinner and free time June 29: 8:00- 10:00 Trad-MCN slide X-ray treated 19:00 – 21:00 Lectures (2 speakers)•Radioactivity, Radiation source, External Radiation intensity, Inverse Square Law- distance/intensity relationship, Exposure dosage, measurement.2) Pollen sterility as the pollution maker (inflos – MCN count. 10:00 – 10:30 Coffee break QA session , Discussion 10:30 - 12:00 Trad-SHM, Average number of hairs per flower, 2nd day mutation scoring. Score sheet. 12:00 – 13:00 Lunch 13:00 – 15:00 Allium root slide preparation of X-ray treated roots 15:00 – 15:30 Coffee break, QA session , Discussion 15:30 – 16:00 Proper use of light microscope, Scoring of MCN. 16:00 – 17:00 Demonstration of SCE assay (full title needed) 17:00-19:00 Dinner and free time 19:00 – 22:00 Lectures and demonstration (3) 1) Radioactive decay, Physical half-life, plotting t1/2 with semi-log paper. Shielding for protection, Half Value Layer 2) Lecture or demonstration 3) Lecture or demonstration June 30: 8:00 -10:00 Trad-MCN slide preparation for proper tetrad stage. Overdose effect, abnormal tetrads, Dis-synchrony of tetrad stages. 10:00 – 10-30 Coffee break, QA session, Discussion 10:30 – 12:00 Trad-MCN slide preparation Trad-SHM assay 3rd day mutation scoring 12:00 – 13:00 Lunch 13:00 - 15:00 Allium slide preparation, Permanent slide prep. 15;00 – 15:30 Coffee break, QA session 15:30 - 18:00 Permanent slide preparation. Statistic Analysis of data- ANOVA, Dunett t statistic 18:00 – 19:00 Banquet for participants and staff together. 19:00 – 22:00 Concluding remarks and Farewell Party

Hands-on Workshop on Plant Genotoxicity Assays – Vienna, Austria. Satellite Program of European Environmental Mutagen Society Meeting–Prague, Czech Republic June 26 -30, 206 June 26: Preparation of plant materials-Tradescantia cuttings, Allium root germination Radioactive source calibration. Audio/Video equipments for evening lectures or demonstrations Treatment of Tradescantia cuttings. with chemical agents,Gaseous, liquid and radiation treat for Trad-MCN assay. Trad cuttings to be treated a week ahead for Trad-SHM assay (S02) June 27: Preparation of plant materials, Laboratory equipment, Dissecting needles stains, reagents (Ethanol, Acetic acid, Carmine stains, hydrochloric acid, vegetable oil) ,Score sheets, lens paper, blotting paper. Registration, lodging accommodation and meal arrangement. June 28: 8:00 – 8:30 Opening ceremony, Power point presentation of 3 (Trad-MCN, SHM, Allium MCN) assays. 8:30- 10:30 Slide preparation - Trad-MCN,Trad-SHM, Allium–root tip, Demonstration, Technical exercise, observation of Micronuclei, pink mutations in stamen hairs, Hair counting, average number of hairs per flower, scoring of mutation frequencies. 10:30 – 10:45 Coffee break and QA session 10:45 – 12:00 Trad-MCN slide preparation. Trad-SHM assay, 1st day Mutation scoring 12:00 – 13:00 Lunch 13:00 – 15:00 Slide prep. meiotic stages in Trad. inflorescences (inflos) 15:00 – 15:30 Coffee break, QA session, Discussion 15:30 – 17:00 Identify meiotic stages in relation to the flower bud sizes. 17:00 – 19:00 Dinner and free time 19:00 – 21:00 Lectures (2 speakers) 1) Radioactivity, Radiation source, External Radiation intensity, Inverse Square Law- distance/intensity relationship, Exposure dosage, measurement. 2) Pollen sterility as the pollution maker (please give a proper title) June 29: 8:00- 10:00 Trad-MCN slide X-ray treated inflos – MCN count. 10:00 – 10:30 Coffee break QA session , Discussion 10:30 - 12:00 Trad-SHM, Average number of hairs per flower, 2nd day mutation scoring. Score sheet. 12:00 – 13:00 Lunch 13:00 – 15:00 Allium root slide preparation of X-ray treated roots 15:00 – 15:30 Coffee break, QA session , Discussion 15:30 – 16:00 Proper use of light microscope, Scoring of MCN. 16:00 – 17:00 Demonstration of SCE assay (full title needed) 17:00-19:00 Dinner and free time 19:00 – 22:00 Lectures and demonstration (3) 1) Radioactive decay, Physical half-life, plotting t1/2 with semi-log paper. Shielding for protection, Half Value Layer 2) Lecture and demonstration 3) Lecture and demonstration June 30: 8:00 -10:00 Trad-MCN slide preparation for proper tetrad stage. Overdose effect, abnormal tetrads, Dis-synchrony of tetrad stages. 10:00 – 10-30 Coffee break, QA session, Discussion 10:30 – 12:00 Trad-MCN slide preparation Trad-SHM assay 3rd day mutation scoring 12:00 – 13:00 Lunch 13:00 - 15:00 Allium slide preparation, Permanent slide prep. 15;00 – 15:30 Coffee break, QA session 15:30 - 18:00 Permanent slide preparation Statistic Analysis of data- ANOVA, Dunett t statistic 18:00 – 19:00 Banquet for participants and staff together. 19:00 – 22:00 Concluding remarks and Farewell Party