Krylov V, Tlapáková T, Mácha J, Curlej J, Ryban L, Chrenek P
Charles University in Prague, Faculty of Science, Department of Cell Biology, Prague, Czech Republic.
Folia Biol (Praha). 2008;54(4):121-4.
For chromosomal localization of the hFVIII human transgene in F2 and F3 generation of transgenic rabbits, FISH-TSA was applied. A short cDNA probe (1250 bp) targeted chromosomes 3, 7, 8, 9 and 18 of an F2 male (animal 1-3-8). Two transgenic offspring (F3) revealed signal positions in chromosome 3 and chromosomes 3 and 7, respectively. Sequencing and structure analysis of the rabbit orthologous gene revealed high similarity to its human counterpart. Part of the sequenced cDNA (1310 bp) served as a probe for FISH-TSA analysis. The rabbit gene was localized in the q arm terminus of the X chromosome. This result is in agreement with reciprocal chromosome painting between the rabbit and the human. The presented FISH-TSA method provides strong signals without any interspecies reactivity.
为了在转基因兔的F2和F3代中对人FVIII转基因进行染色体定位，应用了荧光原位杂交-酪酰胺信号放大（FISH-TSA）技术。一个短的cDNA探针（1250 bp）靶向F2雄性兔（动物1-3-8）的3号、7号、8号、9号和18号染色体。两只转基因后代（F3）分别在3号染色体以及3号和7号染色体上显示出信号位置。兔直系同源基因的测序和结构分析表明其与人类对应基因高度相似。部分测序的cDNA（1310 bp）用作FISH-TSA分析的探针。兔基因定位于X染色体长臂末端。这一结果与兔和人之间的相互染色体涂染结果一致。所展示的FISH-TSA方法提供了强烈的信号，且没有任何种间反应性。
