Reduced body growth and excessive incisor length in insertional mutants
mapping to mouse Chromosome 13
r Tierzucht und Genetik, Veterina
t Wien, Veterina
rplatz 1, A-1210 Wien, Austria
Abteilung Humanbiologie und -genetik, Universita
t Kaiserslautern, Germany
Abteilung Biotechnologie in der Tierproduktion, IFA Tulln, Austria
r Immuno-, Zyto -und Molekulargenetische Forschung, Wien, Austria
Received: 16 January 2002 / Accepted: 2 May 2002
Abstract. Phenotypic and molecular genetic examinations of a
transgenic mouse line showing developmental defects caused
by a recessive insertional mutation were carried out. The
mutant phenotype is characterized by general retardation of
postnatal body growth and by the appearance of increased
incisor length in the upper and lower jaw. The mutation
causing the aberrant phenotype was mapped to Chromosome
13, 40 cM. Examination of the expression of the candidate
genes did not show any alterations. This mutant mouse line
provides a reproducible model for the identi®cation and ex-
amination of gene(s) involved in growth and in the craniofacial
development, including that of the jaws and teeth.
Insertional mutations leading to an abnormal phenotype are
caused by the functional disruption of genes owing to the
random integration of foreign DNA into the host genome.
They have often been found in sites already known to be al-
tered in spontaneous and/or provoked mutants. The appear-
ance of a new phenotype refers to an alteration in the host
genome not previously described or to an unknown eect of a
dierent genetic background on a mutation already described.
Usually, the mutated gene(s) is located at the transgene in-
sertion site. However, in some insertional mutants the abnor-
mal phenotype is caused by secondary alterations mapping to
a distinct locus in the host genome (Rijkers et al. 1994; Meisler
et al. 1998).
Here we describe the phenotypic and molecular genetic
examination of a recessive insertional mutation leading to re-
duced body growth and increased incisor length. Develop-
mental defects resulting in dwar®sm of the aected individuals
are known to be caused by various genetic alterations (Bur-
rows et al. 1999; Conlon and Ra 1999). Knowledge of the
dierent signaling pathways involved in the regulation of tooth
development was obtained by mutant mouse models, mostly
showing the lack of eruption of some or all teeth (Peters and
Balling 1999; Jernvall and Thesle 2000).
Cyclin D1 (Ccnd1)-de®cient mice show a similar phenotype
as observed in our insertional mutants (Fantl et al. 1995; Sic-
inski et al. 1995; Geng et al. 2001). However, reduced body
growth was followed only inconsistently by an increased length
of the incisors. Malformation of the jaw has been suggested as
the cause of the increased incisor length (Fantl et al. 1995).
Ccnd1 maps to mouse Chromosome (Chr) 7 (Mouse Locus
In our insertional mutants, we observed close linkage of the
mutation responsible for the abnormal phenotype and the
transgene integration site. After having mapped the transgene
integration site to a de®ned region of Chr 13, we observed the
presence of the expression of Ccnd1 as well as of selected can-
didate genes known to map to the aected chromosomal site.
The gene responsible for the mutation is yet to be identi®ed.
Materials and methods
Generation and analysis of transgenic mice.
Transgenic mice were
produced by using the gene construct ptrTyr4 and NMRI outbred
mice as described previously (Aigner and Brem 1994). The transgene
encodes a functional mouse tyrosinase and has a length of 15.5 kb.
Direct DNA microinjection into the pronuclei of fertilized oocytes was
carried out according to standard protocols, resulting in the generation
of 11 independent transgenic lines. Molecular genetic analysis of the
transgenic mice was carried out as described (Aigner et al. 1999).
After hemizygous transgenic mice were crossed, homozygous
transgenic ospring of the line NMRI-Tg(Tyr)6 showed an abnormal
phenotype. The ten other tyrosinase transgenic lines did not show the
unexpected phenotype. Integration of about 50 gene construct copies
per hemizygous transgenic cell was observed (Aigner and Brem 1994).
For this study, transgenic mice of generation F20 to F25 were used
after the stable, long-term germline transmission of the transgene in-
tegration site was proven in the line. Molecular genetic dierentiation
of homozygous transgenic and hemizygous transgenic animals was
carried out by Southern analysis, resulting in dierent strengths of the
line-speci®c transgene signal pattern. The molecular genetic analysis
completely con®rmed the strict copy number-dependent expression of
the integrated transgene constructs in the line NMRI-Tg(Tyr)6, which
led to the phenotypically detectable dierence in the intensity of coat
pigmentation in homozygous transgenic and hemizygous transgenic
littermates (Aigner et al. 1999).
Morphologic examination of the insertional mutants.
growth was examined in ospring after mating of hemizygous trans-
genic animals. At birth, the numbers of transgenic and non-transgenic
ospring were recorded from the presence or absence of eye pigmen-
tation, respectively. Body weight was recorded daily from day 1 post
partum (p.p.) to the time point of weaning, 3 weeks p.p. Dierentiation
of homozygous transgenic and hemizygous transgenic mice was carried
out by phenotypic analysis of the coat color intensity and was subse-
quently con®rmed by Southern analysis in randomly selected litters.
The eect of the genetic background on the abnormal phenotype
observed in homozygous transgenic NMRI-Tg(Tyr)6 mice was ex-
amined by backcross experiments (hemizygous transgenic ´ wild type).
Mammalian Genome 13, 504±509 (2002).
Correspondence to: B. Aigner; e-mail: email@example.com-