家族性肌萎缩侧索硬化症转基因鼠的繁殖和鉴定

Establishment of transgenic mouse model of familial amyotrophic lateral sclerosis and identification of the filial generation

目的建立家族性肌萎缩侧索硬化症(FALS)转基因模型鼠的繁育方法,并对其子代鼠进行基因鉴定。方法(1)以6只B6SJL SOD1G93A/+半合子雄鼠与6只B6SJLF1/J+/+雌鼠(1∶1)交配;(2)由鼠尾血提取基因组DNA,PCR扩增hmSOD1基因的片段,电泳后观察结果;(3)对PCR产物进行纯化测序,并通过BLAST验证。结果6对种鼠交配产鼠仔53只,存活率为98%(52/53);G93A hmSOD1阳性鼠约占44.2%(23/52);PCR扩增产物分别为内对照(IL-2):324 bp;Tg(hmSOD1):236 bp;测序证实PCR产物的基因序列和hmSOD1基因片段中的序列一致,并存在G93A突变。结论B6SJL-Tg(SOD1-G93A)1Gur/J雄鼠与B6SJLF1/J雌鼠配种能成功繁育出Tg(hmSOD1)阳性的ALS半合子子代鼠;本实验的PCR法能准确鉴定hmSOD1基因阳性鼠,并证实该转入的基因按近似孟德尔方式遗传,为ALS实验研究奠定了基础。

Objective To establish transgenic mouse models of familial amyotrophic lateral sclerosis （FALS） and identify the genotype of the first filial generation. Methods Six male B6SJL SODIG93A/＋ hemizygote mice were mated with 6 female B6SJLFI/J＋/＋ mice to produce the filial generation. The genomic DNA was extracted from the tail vein blood of the first filial generation mice and PCR was performed to amplify the hmSODl gene fragment. The genotype of the mice was determined by electrophoresis, and the PCR product was purified for further gene sequence analysis and detection of mutation loci. Results Fifty-three progeny mice were born and the survival rate before ALS onset was 98% （52/53）, and among the survived mice, the positivity rate for hmSODl gene was 44.2 % （23/52）. Electrophoresis result showed that the PCR product of 236 bp was consistent with the hmSODl gene fragment, and the sequence of the PCR product was identical with hmSODl gene sequence of G93A mutant. Conclusion Transgenic mouse models of ALS can be established in the first filial generation of male B6SJL SOD 1G93A/＋ mice mated with female B6SJLF 1/J＋/＋. PCR technique can precisely identify the genotype of the filial generation.