用胚泡注射法产生小鼠嵌合体

Production of Mouse Chimeras by Blastocyst Injection

<正> 用免疫手术法分离小鼠胚泡的内细胞团,然后通过显微操作把内细胞团注入另一品系小鼠的胚泡腔内,经体外培养和移植到受体,生下了嵌合体小鼠.其方法是将淡棕色小鼠[(昆明白×C_(57))杂种自交20代以上后培育成的]或昆明白小鼠的胚泡置于0.5%链霉旦白酶中去透明带,然后把裸胚放入1:4稀释的兔抗鼠血清中去滋胚层。抗血清是连续5周。

In present study,mouse chimeras were produced by injecting the inner cell mass(ICM),isolated by immunosurgery,into blastocoele of a mouse blastocyst obtained from the other strain,then cultured and transfered to recipient.The mouse blastocysts of(Kong Min×C_(57))F_1 inbred line(over twenty generations)with light brown colour or Kong Min strain mice with white colour were incubated in 0.5% pronase for removing zonae pellucidae.Zona-free blastocysts were then incubated in 1:4 dilution of rabbit anti-mouse antiserum,which was produced by injection of mouse splenocytes into rabbit vein,for removing trophectoderm.The antiserum was titered and harvested from arteria carotis.The recipient embryo was held by a microholdibg pipette with an outer diameter of 100μm,which was seted on Leitz micromanipula- tar.Isolated inner cell mass was intreduced by puncturing the injection pipette with an outer diameter of 30μm throng zona pellucida and trophectoderm into the blastocoele of the recipient blastocyst.The blasto- cysts were cultured overninht to examine whether it survivaled or not.137 blastocysts were injected and 121 survivaled.Survival rate is 88%.98 blastocysts were transferred into 9 uterine of synchronic pseudo- pregnant recipient females.6 mice were diagnosed as being pregnant and five recipients gave birth to 22 offsprings,pregnant rate was 66% 12 offsprings were devoured by the recipients or foster mother,2 off- springs judged by observation of hair were chimeras in 6 haired in 10 servivaled.The rate of chimerism was 33%.The cell and tissue or other genetic markers will be checked in the near future.