Schedl A, Beermann F, Thies E, Montoliu L, Kelsey G, Schütz G
Institute of Cell and Tumor Biology, German Cancer Research Center, Heidelberg.
Nucleic Acids Res. 1992 Jun 25;20(12):3073-7. doi: 10.1093/nar/20.12.3073.
Transgenic mice have become invaluable for analysing gene function and regulation in vivo. However, the size of constructs injected has been limited by the cloning capacity of conventional vectors, a constraint that could be overcome with yeast artificial chromosomes (YACs). We investigated the feasibility of making transgenic mice with YACs by pronuclear injection of a small YAC carrying a gene encoding tyrosinase. Use of a vector with a conditional centromere allowed fifteenfold amplification of the YAC in yeast and its recovery in high yield. The albino phenotype of the recipient mice was rescued demonstrating the correct expression of the tyrosine gene from the construct. Furthermore, the telomeric sequences added by the yeast integrated into the mouse genome and did not reduce efficiency of integration. Using this technique future experiments with longer YACs will allow the expression of gene complexes such as Hox and the globin gene clusters to be analysed in transgenic animals.
转基因小鼠对于体内基因功能和调控的分析已变得至关重要。然而，注入构建体的大小受到传统载体克隆能力的限制，而酵母人工染色体（YACs）可以克服这一限制。我们通过原核注射携带编码酪氨酸酶基因的小YAC，研究了用YAC制作转基因小鼠的可行性。使用带有条件着丝粒的载体可使YAC在酵母中扩增15倍并高产回收。受体小鼠的白化病表型得到挽救，证明构建体中酪氨酸基因的正确表达。此外，酵母添加的端粒序列整合到小鼠基因组中，且并未降低整合效率。利用这项技术，未来使用更长YACs的实验将能够在转基因动物中分析诸如Hox和珠蛋白基因簇等基因复合体的表达。
