Vadolas Jim, Wardan Hady, Bosmans Marco, Zaibak Faten, Jamsai Duangporn, Voullaire Lucille, Williamson Robert, Ioannou Panos A
Cell and Gene Therapy Research Group, The Murdoch Childrens Research Institute, The University of Melbourne, Royal Children's Hospital, Flemington Road, Parkville, 3052 Melbourne, Australia.
Biochim Biophys Acta. 2005 May 1;1728(3):150-62. doi: 10.1016/j.bbaexp.2005.02.008. Epub 2005 Mar 17.
We report the generation and characterisation of the first transgenic mice exclusively expressing normal human beta-globin ((hu)beta-globin) from a 183 kb genomic fragment. Four independent lines were generated, each containing 2-6 copies of the (hu)beta-globin locus at a single integration site. Steady state levels of (hu)beta-globin protein were dependent on transgene copy number, but independent of the site of integration. Hemizygosity for the transgene on a heterozygous knockout background ((hu)beta(+/0), (mu)beta(th-3/+)) complemented fully the hematological abnormalities associated with the heterozygous knockout mutation in all four lines. Importantly, the rescue of the embryonic lethal phenotype that is characteristic of homozygosity for the knockout mutation was also demonstrated in two transgenic lines that were homozygous for two copies of the (hu)beta-globin locus, and in one transgenic line, which was hemizygous for six copies of the (hu)beta-globin locus. Our results illustrate the importance of transgene copy number determination and of the hemizygosity/homozygosity status in phenotypic complementation studies of transgenic mice containing large heterologous transgenes. Transgenic mouse colonies with 100% (hu)beta-globin production from the intact (hu)beta-globin locus have been established and will be invaluable in comparative and gene therapy studies with mouse models containing specific beta-thalassemia mutations in the (hu)beta-globin locus.
我们报道了首例从183 kb基因组片段中专门表达正常人β-珠蛋白（(hu)β-珠蛋白）的转基因小鼠的产生及特性研究。我们获得了4个独立的品系，每个品系在单个整合位点含有2 - 6个(hu)β-珠蛋白基因座拷贝。(hu)β-珠蛋白的稳态水平取决于转基因拷贝数，但与整合位点无关。在杂合敲除背景（(hu)β(+/0)，(mu)β(th - 3/+)）下，转基因半合子在所有4个品系中完全弥补了与杂合敲除突变相关的血液学异常。重要的是，在两个含有两个拷贝(hu)β-珠蛋白基因座纯合子的转基因品系以及一个含有六个拷贝(hu)β-珠蛋白基因座半合子的转基因品系中，也证明了对敲除突变纯合子特征性的胚胎致死表型的挽救。我们的结果说明了转基因拷贝数测定以及半合子/纯合子状态在含有大型异源转基因的转基因小鼠表型互补研究中的重要性。已建立了从完整(hu)β-珠蛋白基因座产生100% (hu)β-珠蛋白的转基因小鼠群体，这对于与在(hu)β-珠蛋白基因座含有特定β-地中海贫血突变的小鼠模型进行比较研究和基因治疗研究将具有巨大价值。
