Localization of Human Coagulation Factor VIII (hFVIII) in Transgenic Rabbit by FISH-TSA: Identification of Transgene Copy Number and Transmission to the Next Generation

Krylov, Vladimir; Tlapáková, T.; Mácha, J.; Curlej, J.; Ryban, L.; Chrenek, P.

doi:10.14712/fb2008054040121

Folia Biologica > Archive > 2008, Vol. 54 > Issue 4 > Localization of Human Coagulation Factor…

Fol. Biol. 2008, 54, 121-124

https://doi.org/10.14712/fb2008054040121

Localization of Human Coagulation Factor VIII (hFVIII) in Transgenic Rabbit by FISH-TSA: Identification of Transgene Copy Number and Transmission to the Next Generation

Vladimir Krylov¹, T. Tlapáková¹, J. Mácha¹, J. Curlej^2,3, L. Ryban⁴, P. Chrenek^3,5

¹Charles University in Prague, Faculty of Science, Department of Cell Biology, Prague, Czech Republic
²Faculty of Biotechnology and Food Science, Slovak Agricultural University, Nitra, Slovak Republic
³Slovak Agricultural Research Centre, Research Institute of Animal Production, Department of Genetics and Animal Reproduction, Nitra, Slovak Republic
⁴Department of Vascular Biology and Thrombosis Research, Medical University of Vienna, Vienna, Austria
⁵Department of Zoology and Anthropology, Constantine the Philosopher University, Nitra, Slovak Republic

Received June 2008
Accepted August 2008

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 off-spring (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.

Keywords

rabbit, hFVIII, transgenesis, FISH-TSA.

Funding

This work was supported by the Slovak Research and Development Agency (Contract No.: LPP-0126-06) and by the Ministry of Education, Youth and Sports of the Czech Republic (Grant No.: MSM0021620858).