去泛素化酶OTUD6B截短体小鼠模型构建及表型初步分析

Construction of a deubiquitinase OTUD6B truncation mouse model and analysis of phenotype

目的 通过构建去泛素化酶OTUD6B氨基端截短缺失小鼠模型,初步分析OTUD6B氨基端CC2结构域缺失对小鼠发育的影响。方法 利用Cre-Loxp原理,将OTUD6B基因中编码CC2结构域的序列选为打靶序列,构建OTUD6B氨基端截短缺失小鼠模型。利用PCR和Western印迹检测OTUD6B的表达水平,对子代基因型小鼠进行数量统计,利用苏木素-伊红(HE)染色对各脏器进行病理学分析。结果 OTUD6B截短体小鼠模型建立成功。杂合子自交后代小鼠各基因型比例符合孟德尔定律,提示截短体小鼠非胚胎致死,但部分纯合子死于出生后12~48 h。与野生型相比,截短体小鼠的体重较轻、体型较小,主动脉和肺动脉管壁较薄,心肌细胞排列疏松。脾、肺和肝有出血倾向,肾小管上皮细胞呈空泡样变性,其他组织器官无明显差异。结论 成功构建OTUD6B截短体小鼠模型,部分纯合子在出生后12~48 h死亡,死亡小鼠的心、脾、肺、肾等器官出现病变。

Objective To establish a model of OTUD6B truncation mice and analyze the effects on mouse development.Methods The CC2 domain was set as the target sequence and the two Loxp sequences were inserted at the opposite ends of the CC2 domain using Cre-Loxp technology. Then,the genotype was verified by PCR,and the expression of OTUD6B in various tissues was detected by Western blotting to verify the model establishment of truncation mice. Mathematical statistical analysis was conducted to determine whether homozygous OTUD6B mice were born at the expected Mendelian frequency. Hematoxylin-eosin(H&E)staining was used to analyze the pathological changes in all tissues and organs of the offspring. Results Genotype identification and Western blotting analysis demonstrated that an OTUD6B truncation mouse model was established. The percentage of genotypes of mice after self-inbreeding conformed to Mendel′s law,suggesting that OTUD6B truncation mice were not embryonically leathal. However,most of the homozygous mice died in 12-48 h after birth. The body size and mass of truncation mice were smaller than those of the wild-type mice in the same litter. Heart dysgenesis developed in truncation mice,and hemorrhagic lesions developed in the spleen,lung and liver. There were no obvious pathological abnormalities in other tissues and organs. Conclusion An OTUD6B gene truncation mouse model is generated. Most of the homozygous truncation mice die in 12-48 h after birth. Some tissues and organs of dead mutant mice develop lesions,including the heart,spleen,lung and kidney.