Objective: To determine the feasibility of using pUC 118NX plasmid-based transgenic mice in the study of induced mutation in vivo with known mutagen, ethylnitrosourea (ENU).
Methods: pUC 118NX plasmid was recovered from the spleen genomic DNA of ENU-treated and untreated xylE, C57BL/6J transgenic mice with enzyme digestion and circularizing. The recovered pUC 118NX plasmid was electroporated into DH10B host cells, which were incubated in LB solid medium containing proper ampicillin overnight and then sprayed with catechol solution. Mutant could be detected by difference in yellow and white color of the stains and its xylE target gene could be sequenced for further identifying its mutation type. In addition, peripheral blood and bone marrow cells were isolated from xylE, C57BL/6J transgenic mice and micronucleus frequency (MNF) induced by ENU was observed.
Results: The spontaneous mutant frequency for xylE gene in the spleen of xylE, C57BL/6J transgenic mice was less than 4.79 x 10(-5), significantly different from that treated with ENU (50 mg/kg), 19.83 x 10(-5). Types of gene mutation induced by ENU in the spleen of xylE, C57BL/6J transgenic mice included transversion (50%), one or two-base insertion (37.5%) and transition (12.5%). MNF in peripheral blood normochromatic erythrocyte (NCE) and in bone marrow polychromatic erythrocyte (PCE) of ENU-treated (50 mg/kg x 5) mice were 7.6 per thousand and 8.8 per thousand, respectively, both significantly different from the controls treated with solvent (P < 0.01). It indicated that chromosome aberration could be induced by ENU.
Conclusion: The pUC118NX plasmid-based xylE, C57BL/6J transgenic mice, which could be used in detecting gene mutation and chromosome aberration in vivo simultaneously, provided a novel detection system for overall assessment of genetic toxicity of chemicals.