乙基亚硝基脲诱发转基因小鼠体内基因突变及微核形成  

Studies on gene mutation and micronucleus formation induced by ethylnitrosourea (ENU) in transgenic mice in vivo

作　　者：曹洁[1] 印木泉[1] 陈建泉[2] 徐少甫[2] 

机构地区：[1]上海市第二军医大学卫生毒理学教研室,200433 [2]扬州大学农学院动物医学系

出　　处：《中华预防医学杂志》2000年第1期28-30,共3页Chinese Journal of Preventive Medicine

摘　　要：目的　用致突变剂乙基亚硝基脲 (ENU)验证基于pUC 118NX质粒的转基因小鼠用于体内诱变研究的可行性。方法　用酶切、环化方法从经过和未经过ENU诱变处理的xylE ,C5 7BL 6J转基因小鼠脾脏组织DNA中重新分离pUC 118NX质粒 ,使其转化DH10B宿主菌并铺氨苄青霉素抗性平板培养后喷洒邻苯二酚溶液 ,通过菌落颜色不同筛选突变体 ,并对突变靶基因xylE测序分析以确定突变类型。同时取该转基因小鼠外周血及骨髓细胞 ,观察ENU对转基因小鼠微核形成率的影响。结果　转基因小鼠脾脏组织xylE基因自发突变率 <4 79× 10 - 5,经ENU( 5 0mg kg× 5 )处理后 ,脾组织基因诱发突变率为 19 83× 10 - 5,两者差异有显著性。ENU诱发小鼠脾组织基因突变的类型包括颠换( 5 0 % )、单 /双碱基插入 ( 3 7 5 % )和转换 ( 12 5 % )。经ENU( 5 0mg kg× 5 )处理后 ,转基因小鼠外周血细胞微核率 ( 7 6‰ )和骨髓细胞微核率 ( 8 8‰ )均明显高于溶剂对照组 ,表明ENU可诱发该转基因小鼠染色体畸变。结论　基于pUC 118NX质粒的xylE ,C5 7BL 6J转基因小鼠可用于同时测定体内基因突变和染色体畸变 ,有望成为一种新的评价化学物遗传毒性的检测系统。Objective TodeterminethefeasibilityofusingpUC 118NXplasmid basedtransgenicmicein thestudyofinducedmutationinvivowithknownmutagen ,ethylnitrosourea (ENU) .Methods pUC 118NX plasmidwasrecoveredfromthespleengenomicDNAofENU treatedanduntreatedxylE ,C5 7BL 6Jtransgenic micewithenzymedigestionandcircularizing .TherecoveredpUC 118NXplasmidwaselectroporatedintoDH10B hostcells,whichwereincubatedinLBsolidmediumcontainingproperampicillinovernightandthensprayedwith catecholsolution .MutantcouldbedetectedbydifferenceinyellowandwhitecolorofthestainsanditsxylEtarget genecouldbesequencedforfurtheridentifyingitsmutationtype.Inaddition ,peripheralbloodandbonemarrow cellswereisolatedfromxylE ,C5 7BL 6Jtransgenicmiceandmicronucleusfrequency (MNF)inducedbyENUwas observed .Results ThespontaneousmutantfrequencyforxylEgeneinthespleenofxylE ,C5 7BL 6Jtransgenic micewaslessthan 4 79× 10 - 5,significantlydifferentfromthattreatedwithENU ( 5 0mg kg) ,19 83× 10 - 5. TypesofgenemutationinducedbyENUinthespleenofxylE ,C5 7BL 6Jtransgenicmiceincludedtransversion ( 5 0 % )、oneortwo baseinsertion ( 3 7 5 % )andtransition ( 12 5 % ) .MNFinperipheralbloodnormochromatic erythrocyte (NCE)andinbonemarrowpolychromaticerythrocyte (PCE)ofENU treated ( 5 0mg kg× 5 )mice were 7 6‰and 8 8‰ ,respectively ,bothsignificantlydifferentfromthecontrolstreatedwithsolvent (P < 0 0 1) .ItindicatedthatchromosomeaberrationcouldbeinducedbyENU .Conclusion ThepUC118NXplasmid basedxylE ,C5 7BL 6Jtransgenicmice,whichcouldbeusedindetectinggenemutationandchromosome aberrationinvivosimultaneously ,providedanoveldetectionsystemforoverallassessmentofgenetictoxicityof chemicals.

关 键 词：小鼠 转基因 微核 乙基亚硝脲 DNA损伤 

分 类 号：R730.231[医药卫生—肿瘤]