Localization of human coagulation factor VIII (hFVIII) in transgenic rabbit by FISH-TSA: identification of transgene copy number and transmission to the next generation.
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.
Krylov
V
V
Charles University in Prague, Faculty of Science, Department of Cell Biology, Prague, Czech Republic. vkrylov@natur.cuni.cz
Tlapáková
T
T
Mácha
J
J
Curlej
J
J
Ryban
L
L
Chrenek
P
P
D000818:Animals; D030801:Animals, Genetically Modified; D002874:Chromosome Mapping / Q000379:methods*; D033481:Chromosomes, Mammalian; D005169:Factor VIII / Q000235:genetics* / Q000378:metabolism; D005260:Female; D018628:Gene Dosage; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008297:Male; D021141:Nucleic Acid Amplification Techniques; D011817:Rabbits; D019076:Transgenes / Q000235:genetics
D005169:Factor VIII