Broome R L, Feng L, Zhou Q, Smith A, Hahn N, Matsui S M, Omary M B
Department of Veterinary Medicine, VA Palo Alto Health Care System, 3801 Miranda Avenue, 154J, Palo Alto, CA 94304, USA.
FEBS Lett. 1999 Nov 26;462(1-2):159-60. doi: 10.1016/s0014-5793(99)01516-1.
Commonly applied genotyping of transgenic mice involves using tail or ear biopsies which may cause discomfort to the animal. We tested the possibility of polymerase chain reaction (PCR)-based mouse genotyping using stool specimens from three transgenic mouse lines that overexpress 10-18 transgene copies of human keratin polypeptide 18, as compared to genotyping using tail biopsies. Stool specimens were obtained with ease and provided easy detection of the human transgene product. The method was also able to detect endogenous mouse actin and keratin genes which presumably are present at two copies each. Nested PCR was not necessary for genotyping using stool-derived genomic material but did increase the relative magnitude of the signal obtained. The non-invasive genotyping method described herein offers a reproducible, sensitive and effective modality that could replace invasive tissue sampling procedures currently used to test thousands of genetically altered mice.
常用的转基因小鼠基因分型方法包括采集尾巴或耳部活检样本,这可能会给动物带来不适。我们测试了基于聚合酶链反应(PCR)的小鼠基因分型方法,该方法使用来自三个转基因小鼠品系的粪便样本进行基因分型,这些品系分别过量表达10 - 18个转基因拷贝的人角蛋白多肽18,并将其与使用尾巴活检样本进行基因分型的方法进行了比较。粪便样本很容易获得,并且能够轻松检测到人转基因产物。该方法还能够检测内源性小鼠肌动蛋白和角蛋白基因,推测每个基因各有两个拷贝。使用粪便来源的基因组材料进行基因分型时,不需要巢式PCR,但它确实增加了获得的信号的相对强度。本文所述的非侵入性基因分型方法提供了一种可重复、灵敏且有效的方式,可以取代目前用于检测数千只基因改造小鼠的侵入性组织采样程序。