title: Instability of the insertional mutation in CftrTgH(neoim)Hgu cystic fibrosis mouse model.
abstract: BACKGROUND
  A major boost to the cystic fibrosis disease research was given by the generation of various mouse models using gene targeting in embryonal stem cells. Moreover, the introduction of the same mutation on different inbred strains generating congenic strains facilitated the search for modifier genes. From the original CftrTgH(neoim)Hgu CF mouse model we have generated using strict brother x sister mating two inbred CftrTgH(neoim)Hgu mouse lines (CF/1 and CF/3). Thereafter, the insertional mutation was introgressed from CF/3 into three inbred backgrounds (C57BL/6, BALB/c, DBA/2J) generating congenic animals. In every backcross cycle germline transmission of the insertional mutation was monitored by direct probing the insertion via Southern RFLP. In order to bypass this time consuming procedure we devised an alternative PCR based protocol whereby mouse strains are differentiated at the Cftr locus by Cftr intragenic microsatellite genotypes that are tightly linked to the disrupted locus.
  
  
  RESULTS
  Using this method we were able to identify animals carrying the insertional mutation based upon the differential haplotypic backgrounds of the three inbred strains and the mutant CftrTgH(neoim)Hgu at the Cftr locus. Moreover, this method facilitated the identification of the precise vector excision from the disrupted Cftr locus in two out of 57 typed animals. This reversion to wild type status took place without any loss of sequence revealing the instability of insertional mutations during the production of congenic animals.
  
  
  CONCLUSIONS
  We present intragenic microsatellite markers as a tool for fast and efficient identification of the introgressed locus of interest in the recipient strain during congenic animal breeding. Moreover, the same genotyping method allowed the identification of a vector excision event, posing questions on the stability of insertional mutations in mice.
authors.affiliation: Klinische Forschergruppe, OE 6710, Medizinische Hochschule Hannover, D-30625 Hannover, Germany. charizopoulou.nikoletta@mh-hannover.de
authors.forename: Nikoletta
authors.initials: N
authors.lastname: Charizopoulou
authors.forename: Silke
authors.initials: S
authors.lastname: Jansen
authors.forename: Martina
authors.initials: M
authors.lastname: Dorsch
authors.forename: Frauke
authors.initials: F
authors.lastname: Stanke
authors.forename: Julia R
authors.initials: JR
authors.lastname: Dorin
authors.forename: Hans-Jürgen
authors.initials: HJ
authors.lastname: Hedrich
authors.forename: Burkhard
authors.initials: B
authors.lastname: Tümmler
mesh_terms: D000818:Animals; D003433:Crosses, Genetic; D003550:Cystic Fibrosis / Q000235:genetics*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D004195:Disease Models, Animal; D005260:Female; D005787:Gene Frequency; D005838:Genotype; D018095:Germ-Line Mutation / Q000235:genetics; D006239:Haplotypes / Q000235:genetics; D008297:Male; D051379:Mice; D020297:Mice, Congenic; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D018895:Microsatellite Repeats; D016254:Mutagenesis, Insertional; D009154:Mutation / Q000235:genetics*
chemical_list: D019005:Cystic Fibrosis Transmembrane Conductance Regulator