DEVELOPMENT OF THE SYSTEM FOR CREATING A TRANSGENIC MINIPIG WITH THE THREE-DIMENSIONAL MONITORING SYSTEM

本项目的目的是建立一个高效的用于异种器官移植的转基因小型猪系统。我们发明了三维（3D）刚性内窥镜，用于在不进行剖腹手术的情况下从卵巢获得微创小型猪卵子，以及具有由微孔玻璃板制成的5 X 5微袋基质的卵子保持装置。该装置可以将卵子吸入直径为150微米的微袋中，溶液在固定在保持器壳体中的板上循环。卵可以在适当的温度、pH和气体含量的受控环境下在装置中培养。GFP标记的人基因可以通过计算机引导的显微注射器导入细胞核。所有的过程都可以用3D显微镜监控，并用录像机记录下来。培养的GFP标记的胚胎可以被选择性地拾取并用3D刚性内窥镜低侵入性地通过腹腔镜引入雌性小型猪子宫中。该系统可以比传统方法更有效地创建转基因小型猪几十倍，传统方法需要更多的技术和更多的雌性小型猪。

Aim of the project is to establish an efficient system of creating transgenic minipigs for heterogenic organ transplantation. We invented three-dimensional (3D) rigid endoscopes for obtaining less-invasively minipig ova from ovarium without laparotomy and an ova-holding device having 5 x 5 micropocket matrix made of microporous glass plate. The device can suck ova into micropockets having diameter of 150 micrometer with solution circulating across the plate fixed in the holder housing. Ova can be cultured in the device under controlled environments of appriate temperature, pH, and gas contents. GFP marked-human gene can be introduced into the nucleus with the computer guided microinjector. All the procedures can be monitored with the 3 D microscope and recorded with a video-cassette recorder. The cultured GFP- marked embryo can be selectively picked up and introduced laparoscopically into female minipig uterus less-invasively with the 3 D rigid endoscope. This system can create several ten times more efficiently a trangenic minipig than the conventional method which needs much skill and more female minipig sacrification.