Identification of DNA polymorphism induced by X-ray and UV irradiation in plant cells

Angelonia protoplasts were irradiated with X-rays or UV to obtain a half survival rate. To evaluate the DNA damage, total genomic DNA was prepared from protoplasts irradiated with X-rays or UV and nonirradiated control protoplasts, respectively. An aliquot of DNA was subjected to polymerase chain reaction (PCR) with a decamer arbitrary primer in the presence of radiolabeled nucleotides under annealing conditions of low stringency. Randomly amplified PCR products were analyzed using a denaturing polyacrylamide gel. All the DNA bands detected by autoradiography were compared. The number of DNA bands detected in one lane was approximately similar to 150. The results indicate that the arbitrarily amplified PCR method is very sensitive for monitoring the DNA polymorphism caused by X-ray and UV irradiation. This novel procedure, designated as RI (radioisotope incorporated)-RAPD, could be applied to detect precise DNA polymorphism in plant cells.