绿色荧光蛋白转基因小鼠用作hprt基因突变试验动物模型的研究

Green fluorescent protein transgenic mice as an animal model for hprt gene mutation test

目的:探讨绿色荧光蛋白(green fluorescent protein,GFP)转基因小鼠用作hprt基因突变试验动物模型的可能性。方法:用乙基亚硝基脲(N-ethyl-N-nitrosourea,ENU)诱导GFP转基因小鼠和普通小鼠基因突变,比较诱导后两种小鼠的嗜多染红细胞微核率变化情况。再观察GFP小鼠细胞克隆对GFP转基因小鼠hprt基因的DNA和mRNA序列进行验证,并对GFP转基因小鼠hprt突变克隆外显子1和外显子9进行了双向测序。结果:ENU诱导后GFP转基因小鼠和普通小鼠的微核率变化趋势相同。GFP转基因小鼠的微孔培养物比普通小鼠的微孔培养物易于确认,且降低了假阳性率和假阴性率。GFP转基因小鼠的hprt基因是完整的,与普通小鼠相比未见明显差异。GFP转基因小鼠hprt基因6个突变克隆外显子1的序列均与野生型小鼠细胞的序列一致,3个突变克隆外显子9的序列中有2个发生了A∶T→T∶A颠换。结论:ENU诱导的GFP转基因小鼠突变模型良好,GFP转基因小鼠可用作小鼠淋巴细胞hprt基因突变试验的动物模型,准确性高且试验难度低。

OBJECTIVE： To investigate whether GFP transgenic mice can be used as a new animal model for hprt gene mutation test. METHODS： ENU was used to induce the gene mutation of GFP transgenic and normal mice. The dynamic of frequencies of micronuclei in peripheral blood PEC was compared between GFP transgenic and normal mice after ENU induction. DNA and mRNA sequences of hprt were tested in GFP transgenic mice. PCR-sequencing was used to detect six exons 1 and three exons 9. RESULTS： The dynamic of frequencies of micronuclei was consistent between GFP transgenic and normal mice after ENU induction. The materials of GFP transgenic mice were easier to confirm than those of normal mice. It reduced the false positive rate and false negative rate as well. The hprt gene sequences of GFP transgenic mice were complete. There were no significant differences in hprt gene sequences between GFP transgenic and normal mice. DNA sequences of exons 1 were consistent with the wild-type mouse. Two A ： T→T ： A substitutions occurred in exons 9 of hprt gene from mutation clone. CONCLUSION ： GFP transgenic mice can be used as animal models for mutation test. GFP transgenic mice can be used as the animal model with high accuracy for hprt gene mutation test, which may reduce the difficulty of the test.