Detection of mitomycin C-induced genetic damage in fish cells by use of RAPD

Abstract

Concern about genetic alterations in fish populations arising from anthropogenic activities has led to the adaptation and/or development of new tests and techniques that shed light on these alterations. The high number and the reduced size of chromosomes and the long cell cycle associated with most fish species preclude the use of most accepted genotoxicity assays. The purpose of this work was to study the capability of the randomly amplified polymorphic DNA technique to show genotoxic effects induced by chemicals in fish cells. To do that we studied the effect of 0.5 microg/ml mitomycin C (MMC) on an established rainbow trout cell line (RTG-2). To increase the sensitivity of detecting altered copies of DNA and to avoid the presence of false positives and a lack of reproducibility, the amounts of DNA template and primer present in amplification reactions were studied and optimized after comparison between the control and exposed fingerprints for 4, 6 and 8 h. Results show that 5 ng of DNA template and 4 pM chosen primer were optimum to show differences between control and exposed cells and to obtain reproducible results. The results obtained, after optimum conditions were established, show that this system could be useful for the assessment of DNA alterations in in vitro genotoxicity studies.

Detection of physiological and genotoxic damages reflecting toxicity in Kalanchoe clones

“…DNA fingerprinting was reported to be an important bio-indicator used in evaluation of toxicants' effects Becerril et al, 1999;Grayson et al, 1999). The pollutant-induced changes in RAPD profiles were used to comparatively analyze the alterations in genomic DNA stability and genotoxic effects with other parameters (Atienzar et al, 2000;Liu et al, 2005).…”

Genotoxicity Testing in Pesticide Safety Evaluation

“…RAPD-PCR has also been successfully utilized in in vitro genotoxicity test. Becerril et al (1999) studied the effect of the well known carcinogen, mitomycin C in RTG-2 fish cell line by RAPD-PCR. The bands obtained were analyzed to show a difference in the banding pattern of control with that of the exposed groups.…”

Detection of DNA alteration in abnormal phenotype of broiler chicken male by random amplified polymorphic DNA (RAPD)

“…The results from this study are in agreement with findings of several researchers who studied capability of RAPD to detect X-ray-induced DNA damage (Shimada and Shima, 1998), and genetic alterations in human tumors (Ambas et al, 1997). In addition, RAPD assay have also used to detect genetic instability in tumors (Luceri et al, 2000) and DNA alterations induced by benzo[a]pyrene (Atienzar et al, 1999;2002b), copper (Atienzar et al, 2001), mitomycin C (Becerril et al, 1999), chrysotile asbestos (Yoshida et al, 2001), UV or X-rays (Atienzar et al, 2000;Jones and Kortenkamp, 2000), and radionuclide exposure (Theodorakis, 2001). Sharma et al (2001) have earlier established the effectiveness of RAPD in detecting polymorphism between chicken populations and their applicability in population studies and in the establishment of genetic relationships among the chicken populations.…”

“…The AP-PCR method was successfully applied to detect x-rayinduced DNA damage (Shimada and Shima, 1998), genetic alterations in human tumors (Ambas et al, 1997) and to evaluate the involvement of a DNA repair gene following genotoxic treatment (Lopez et al, 1999). In addition, the RAPD assay was also used to detect genetic instability in tumors (Luceri et al, 2000) as well as detected DNA alterations induced by benzo[a]pyrene (Atienzar et al, 20002b), copper (Atienzar et al, 2001), mitomycin C (Becerril et al, 1999), 4-n-nonylphenol and 17-β estradiol (Atienzar et al, 2002c), chrysotile asbestos (Yoshida et al, 2001), UV or X-rays (Atienzar et al, 2000;Jones and Kortenkamp, 2000), and radionuclide exposure (Theodorakis, 2001). Becerril et al (1999) have also studied the capability of the randomly amplified polymorphic DNA technique to show genotoxic effects induced by chemicals in fish cells.…”

“…In addition, the RAPD assay was also used to detect genetic instability in tumors (Luceri et al, 2000) as well as detected DNA alterations induced by benzo[a]pyrene (Atienzar et al, 20002b), copper (Atienzar et al, 2001), mitomycin C (Becerril et al, 1999), 4-n-nonylphenol and 17-β estradiol (Atienzar et al, 2002c), chrysotile asbestos (Yoshida et al, 2001), UV or X-rays (Atienzar et al, 2000;Jones and Kortenkamp, 2000), and radionuclide exposure (Theodorakis, 2001). Becerril et al (1999) have also studied the capability of the randomly amplified polymorphic DNA technique to show genotoxic effects induced by chemicals in fish cells. The results obtained after optimum conditions were established, showed that this system could be useful for the assessment of DNA alterations in vitro genotoxicity studies.…”