CAJ | 학술논문

背景乙酰基亚硝基脲(ENU)诱导小鼠基因突变是研究基因功能及建立人类疾病动物模型的一种有效手段. 目的研究ENU诱变获得的1只角膜混浊小鼠角膜组织形态变化,并对其突变基因进行染色体定位.方法选用8～10周龄的C57BL/6J (B6)雄鼠40只,腹腔内注射ENU.将注射后的雄鼠与同品系母鼠交配,在其后代小鼠中筛选眼部突变个体,并将突变个体与同品系小鼠配种以确定是否遗传.对ENU诱变获得1只角膜混浊小鼠的角膜行组织病理学观察.繁殖[(B6×D2) F1×B6] N2角膜混浊小鼠,用平均分布于小鼠染色体上的微卫星标记对N2小鼠染色体进行扫描.采用优势对数计分法(LOD)判定突变基因与微卫星是否连锁. 结果将角膜呈现混浊小鼠与B6背景小鼠配种,在其59只后代中出现19只类似表型的突变小鼠.角膜组织病理学检查发现,该例角膜混浊小鼠角膜基质增厚,出现新生血管和炎性细胞浸润,成纤维细胞明显增多.突变基因与微卫星间的连锁分析发现,26个N2样品中突变基因与位于小鼠2号染色体63.42 cM处的D2Mi307位点发生3例交换,LOD值为3.79,该突变基因位于小鼠第2号染色体. 结论成功获得角膜混浊小鼠并将其突变基因初步定位于2号染色体,有望为人类角膜混浊疾病研究提供一种新的小鼠模型.
배경 을선기아초기뇨(ENU)유도소서기인돌변시연구기인공능급건립인류질병동물모형적일충유효수단. 목적 연구ENU유변획득적1지각막혼탁소서각막조직형태변화,병대기돌변기인진행염색체정위.방법 선용8～10주령적C57BL/6J (B6)웅서40지,복강내주사ENU.장주사후적웅서여동품계모서교배,재기후대소서중사선안부돌변개체,병장돌변개체여동품계소서배충이학정시부유전.대ENU유변획득1지각막혼탁소서적각막행조직병이학관찰.번식[(B6×D2) F1×B6] N2각막혼탁소서,용평균분포우소서염색체상적미위성표기대N2소서염색체진행소묘.채용우세대수계분법(LOD)판정돌변기인여미위성시부련쇄. 결과 장각막정현혼탁소서여B6배경소서배충,재기59지후대중출현19지유사표형적돌변소서.각막조직병이학검사발현,해례각막혼탁소서각막기질증후,출현신생혈관화염성세포침윤,성섬유세포명현증다.돌변기인여미위성간적련쇄분석발현,26개N2양품중돌변기인여위우소서2호염색체63.42 cM처적D2Mi307위점발생3례교환,LOD치위3.79,해돌변기인위우소서제2호염색체. 결론 성공획득각막혼탁소서병장기돌변기인초보정위우2호염색체,유망위인류각막혼탁질병연구제공일충신적소서모형.
Background N-ethyl-N-nitrosourea (ENU)-induced mouse mutagenesis is a powerful approach for the study of gene function and the generation of human disease models.Objective This study was to create the corneal morphologic change and map the mutant gene of a kind of corneal opacity in ENU mutagenesis in mouse.Methods ENU was intraperitoneally injected in forty C57BL/ 6J (B6) male mice aged 8-10 weeks old.The male mice were mated with the same strain female mice.Their progenies were screened for visible eye mutation,and the mutant mice were mated with the same strain mice to confirm the heredity of mutation phenotypes.Hematoxylin & eosin staining was used to examine the histopathological change of cornea in one mouse with ENU-induced corneal opacity.To map the mutant gene,[(B6×D2)F1 ×B6] N2 mutant mice were bred,and the genome of the N2 mice was scanned by microsatellite markers distributed equally on the mouse chromosome.The microsatellite linked to the mutant gene was determined by the log odds score.This experimental procedure was approved by Ethic Committee about Experimental Animal Care and Use of Yangzhou University.Results The founder mouse,which was the progeny of an ENU-treated B6 male mouse and an untreated B6 female mouse,had a corneal opacity phenotype.After mating the mutant with B6 mice,19 of 59 descendants appeared corneal opacity phenotype.Thickening of corneal stroma,neoangiogenesis,infiltration of inflammatory cells and proliferation of fibroblasts were exhibited in cloudy cornea in ENU-induced mutated mice under the optical microscope.After linkage analysis between microsatellite markers and the mutant gene,the mutant gene was linked to D2Mi307,which was located at 63.42 cM.Three cases of 26 N2 mice underwent recombination with the LOD 3.79.The mutant gene associated with the cornea phenotype was located on chromosome 2.Conclusions This study map the mutant gene associated with the cornea phenotype on chromosome 2.The strain might be used as a mouse model for heritable human corneal opacity.