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靶基因xylE质粒载体转基因小鼠的建立 |
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曹洁 陈建泉 印木泉 成国祥 黄建 毛裕民 徐少甫
【摘要】 目的 建立在基因组中整合有pUC118NX质粒的转基因小鼠致突变检测模型。方法 将含xylE靶基因的pUC118NX质粒DNA显微注射至C57BL/6J小鼠376枚受精卵雄性原核中,存活的225枚受精卵移入11只假孕母鼠双侧输卵管内发育成个体后,用PCR-Southern blot、质粒回收转化试验和酶切鉴定方法筛选存活仔鼠。结果 假孕母鼠中有7只妊娠,共产仔鼠29只, 存活25只，其中PCR-Southern blot检测阳性小鼠18只，阳性率为72％。最后选择2只在基因组中完整整合pUC118NX质粒的健康雄性小鼠作为Founder鼠,进行转基因小鼠致突变检测和建系工作。结论 成功地建立了在基因组DNA中整合有pUC118NX质粒的C57BL/6J转基因小鼠。
【关键词】 小鼠，转基因 质粒 诱变力试验
Development of Plasmid-based Transgenic Mice Carrying with Target Gene xylE CAO Jie*, CHEN Jianquan, YIN Muquan, et al. *Department of Hygienic Toxicology The Second Military Medical University, Shanghai 200433
【Abstract】 Objective To develop a transgenic mouse model carrying with plasmid pUC118NX integrated into its genomic DNA for detecting mutagenesis. Methods DNA of plasmid UC118NX in target gene xy1E was injected microscopically into male protonucleus of 376 mouse spermatova，and 225 survival spermatova were transferred into the oviducts in both sides of 11 pseudopregnant female mice to develop their offspring．The genomic DNA in survival young mice were analyzed with polymerase chain reaction （PCR）-Southern blot， plasmid transformation test and endonuclease-digestion. Results Seven pseudopregnant mice got pregnant， and 29 offspring were delivered and 25 survived of which 18 were identified with positive for PCR-Southern blot（72%）． The two stout male mice with intact integration of plasmid pUC118NX in their genome were finally chosen as founders to detect gene mutation in vivo and establish transgenic mouse lineages. Conclusion Transgenic C57BL/6J mice integrated with plasmid pUC118NX into their genomic DNA have been successfully developed.
【Key words】 Mouse, transgenic Plasmids Mutagenicity tests
自1989年Gossen等［1］建立第1个转基因小鼠突变检测模型以来, 哺乳动物体内基因突变研究才真正成为可能。目前以λ噬菌体为载体的转基因小鼠（如muta mouse和big blue)已开始用于体内基因突变研究和化学物质的遗传毒性分析。但是在这类转基因小鼠模型中，载体的回收需进行体外包装,不仅操作繁琐，且从器官或组织中分离的基因组DNA片段必须很大,保证有足够的cos 位点才能进行[1] [2] 下一页
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