Physical Characterization of the Chromosomal Rearrangements That Accompany the Transgene Insertion in thechakragatiMouse Mutant

The circling phenotype of thechakragatimouse is a result of a transgenic insertional mutation. The absence of the phenotype in mice heterozygous for the transgene insertion suggests that this is due to a loss of function of an endogenous gene. Efforts to identify this gene have led to a previous report that sequences flanking the transgene,D16Ros1andD16Ros2,map 10 cM apart in wildtype mice. We present here physical mapping data indicating that the proximity ofD16Ros1andD16Ros2in theckrmouse is explained by a duplication ofD16Ros2and its insertion along with the transgene atD16Ros1.We further demonstrate thatD16Ros1sequences are also duplicated and that this duplication is also part of the insertion at the endogenousD16Ros1locus.     

Comparative Genome Mapping of the Ataxia–Telangiectasia Region in Mouse, Rat, and Syrian Hamster

Chromosomal locations of theAtm(ataxia–telangiectasia (AT)-mutated) andAcat1(mitochondrial acetoacetyl-CoA thiolase) genes in mouse, rat, and Syrian hamster were determined by direct R-banding FISH. Both genes were colocalized to the C-D band of mouse chromosome 9, the proximal end of q24.1 of rat chromosome 8, and qa4–qa5 of Syrian hamster chromosome 12. The regions in the mouse and rat were homologous to human chromosome 11q. Fine genetic linkage mapping of the mouse AT region was performed using the interspecific backcross mice.Atm, Acat1,andNpat,which is a new gene isolated from the AT region, and 12 flanking microsatellite DNA markers were examined. No recombinations were found among theAtm, Npat, Acat1,andD9Mit6loci, and these loci were mapped 2.0 cM distal toD9Mit99and 1.3 cM proximal toD9Mit102.Comparison of the linkage map of mouse chromosome 9 (MMU9) and that of human chromosome 11 (HSA11) indicates that there is a chromosomal rearrangement due to an inversion betweenEts1andAtm–Npat–Acat1and that the inversion of MMU9 originated from the chromosomal breakage at the boundary betweenGria4andAtm–Npat–Acat1on HSA11. This type of inversion appeared to be conserved in the three rodent species, mouse, rat, and Syrian hamster, using additional comparative mapping data with theRckgene.     

Genetic mapping of two DNA markers,D16Ros1 and D16Ros2, flanking the mutation site in the chakragati mouse,a transgenic insertional mutant

We present here the genetic mapping of two novel loci, D16Ros1 and D16Ros2, to mouse Chromosome (Chr) 16. The probes for these loci were genomic framents isolated from the chakragati mouse, a behavioural mutant resulting from insertional mutagenesis during the course of making transgenic mice. D16Ros1 and D16Ros2 were first mapped by recombinant inbred (RI) strain analysis and subsequently by the analysis of 145 progeny of two interspecific backcrosses between Mus domesticus and Mus spretus. These progeny had been typed for the centromere and this allowed mapping of D16Ros1 and D16Ros2 relative to the centromere. The other markers included in this study were Prm-1, Gap43 and Sod-1. The genetic map generated spanned 47.5 cM from the centromere to Sod-1, the most distal marker mapped here. The linkage data presented here should prove useful in mapping other loci relative to the centromere of Chr 16.     

Fine Genetic Map of Mouse Chromosome 10 around the Polycystic Kidney Disease Gene,jcpk,and Ankyrin 3

A chlorambucil (CHL)-induced mutation of thejcpk(juvenile congenital polycystic kidney disease) gene causes a severe early onset polycystic kidney disease. In an intercross involvingMus musculus castaneus, jcpkwas precisely mapped 0.2 cM distal toD10Mit115and 0.8 cM proximal toD10Mit173.In addition, five genes,Cdc2a, Col6a1, Col6a2, Bcr,andAnk3were mapped in both thisjcpkintercross and a (BALB/c × CAST/Ei)F₁× BALB/c backcross. All five genes were eliminated as possible candidates forjcpkbased on the mapping data. Thejcpkintercross allowed the orientation of theAnk3gene relative to the centromere to be determined.D10Mit115, D10Mit173, D10Mit199,andD10Mit200were separated genetically in this cross. The order and genetic distances of all markers and gene loci mapped in thejcpkintercross were consistent with those derived from the BALB/c backcross, indicating that the CHL-induced lesion has not generated any gross chromosomal abnormalities detectable in these studies.     

Physical and Genetic Maps of thedeafwaddlerRegion on Distal Mouse Chr 6

Thedeafwaddler(dfw) mutation, displaying motor ataxia and profound deafness, arose spontaneously in a C3H/HeJ colony and was mapped previously to distal mouse Chr 6. In this study, a high-resolution genetic map was generated by positioning 10 microsatellite markers and 5 known genes on a 968-meioses intersubspecific backcross segregating fordfw[(CAST/Ei–+/+ × C3HeB/FeJ–dfw/dfw) × C3HeB/FeJ–dfw/dfw], giving the following marker order and sex-averaged distances:D6Mit64–(0.10 + 0.10 cM)–Pang–(1.24 + 0.36 cM)–Itpr1–(0.62 + 0.25 cM)–D6Mit108–(0.52 + 0.23 cM)–D6Mit54–(0.21 + 0.15 cM)–D6Mit23, D6Mit107, D6Mit328–(0.72 + 0.27 cM)–D6Mit11–(0.21 + 0.15 cM)–dfw–(0.93 + 0.31 cM)–Gat4, D6Mit55–(0.10 + 0.10 cM)–D6Mit63–(0.31 + 0.18 cM)–Syn2–(0.62 + 0.25 cM)–D6Mit44(Rho). Female and male genetic maps are similar immediately surrounding thedfwlocus, but show marked differences in other areas. A yeast artificial chromosome-based physical map suggests that the closest markers flanking thedfwlocus,D6Mit11(proximal) andGat4, D6Mit55(distal), are contained within 650–950 kb. The human homologues of the flanking lociItpr1(proximal) andSyn2(distal) map to chromosome 3p25–p26, suggesting that the human homologue of thedfwgene is located within this same region.     

High-Resolution Linkage Map of Mouse Chromosome 13 in the Vicinity of the Host Resistance LocusLgn1

Natural resistance of inbred mouse strains to infection withLegionella pneumophilais controlled by the expression of a single dominant gene on chromosome 13, designatedLgn1.The genetic difference atLgn1is phenotypically expressed as the presence or absence of intracellular replication ofL. pneumophilain host macrophages. In our effort to identify theLgn1gene by positional cloning, we have generated a high-resolution linkage map of theLgn1chromosomal region. For this, we have carried out extensive segregation analysis in a total of 1270 (A/J × C57BL/6J) × A/J informative backcross mice segregating the resistance allele of C57BL/6J and the susceptibility allele of A/J. Additional segregation analyses were carried out in three preexisting panels of C57BL/6J ×Mus spretusinterspecific backcross mice. A total of 39 DNA markers were mapped within an interval of approximately 30 cM overlapping theLgn1region. Combined pedigree analyses for the 5.4-cM segment overlappingLgn1indicated the locus order and the interlocus distances (in cM):D13Mit128–(1.4)–D13Mit194–(0.1)–D13Mit147–(0.9)–D13Mit36–(0.9)–D13Mit146–(0.2)–Lgn1/D13Mit37–(1.0)–D13Mit70.Additional genetic linkage studies of markers not informative in the A/J × C57BL/6J cross positionedD13Mit30, -72, -195,and-203, D13Gor4, D13Hun35,andMtap5in the immediate vicinity of theLgn1locus. The marker density and resolution of this genetic linkage map should allow the construction of a physical map of the region and the isolation of YAC clones overlapping the gene.     

Assignment of the gene for adenine phosphoribosyltransferase on the genetic map of mouse chromosome 8

Two electrophoretic variants of adenine phosphoribosyltransferase (APRT) were identified in a population of wild mice (Mus musculus bactrianus). Breeding tests demonstrated that the APRT variants are under the control of two alleles at an autosomal locus designatedAprt. We have examined the linkage relationships betweenAprt and the markers of chromosome 8 including esterase-1 and the centromere. The recombination distance between the centromere andAprt is 44 ± 7 cM, and that betweenEs-1 andAprt is 25 ± 2 cM, i.e., the probable order of the markers examined is cen-Es-1-Aprt on chromosome 8.     

Molecular organization of theD-Qa region oft-haplotypes suggests that recombination is an important mechanism for generating genetic diversity of the major histocompatibility complex

We have determined the molecular maps of theH-2D andQa regions of thet-complex haplotypest ^l2 andt ^w5 by chromosomal walking. Analysis with class I probes and other probes unique to theH-2D:Qa subregion indicates that the class I gene organization oft ¹² is:D1-D2-Q1-Q2-Q3-Qx-Q4-Q5-Q10, while that oft ^w5 is:D1-D2-Q1-Q2-Q4-Q5-Q10. Thus, the absence of theQ6-Q9 genes suggested previously int-haplotypes was confirmed. A comparison of the molecular maps of thet ¹² andt ^w5 chromosomes revealed an extremely mosaic pattern of diversity: The regions betweenD1 andD2, and betweenQ4 andQ10, are very similar in both chromosomes. However, theirQ1 toQ3 regions are strikingly different. Further comparisons of wild-type chromosomes and additionalt-haplotypes by molecular mapping and genomic Southern blot hybridization with probes to theQ1-Q3 region showed a high level of polymorphism among both wild-type chromosomes and amongt-haplotypes. The characteristics of the polymorphisms suggest that recombination may play an important role in generating this genetic diversity. Furthermore, recombination between wild-type andt-haplotype chromosomes may be involved.     

Genetic mapping of the polycystic kidney gene,pcy, on mouse chromosome 9

The murine polycystic kidney disease gene,pcy, is an autosomal recessive trait located on chromosome 9. To determine the genetic locus ofpcy, 222 intraspecific backcross mice were obtained by mating C57BL/6FG-pcy andMus molossinus. Restriction fragment length polymorphism analysis of 70 of the 222 backcross progeny showed thatpcy, dilute coat color (d), and cholecystokinin (Cck) were located in the orderd—pcy—Cck from the centromere. Simple sequence repeat length polymorphism analysis of DNA of all 222 backcross mice was carried out using four markers which were located near the central regions ofd andCck. One and eight recombinations were detected betweenD9Mit24 andpcy and betweenD9Mit16 andpcy, respectively. However, no recombinant was observed amongpcy, D9Mit14, andD9Mit148. These findings strongly suggest thatD9Mit14 andD9Mit148 are located near thepcy gene and are good markers for chromosomal walking to this gene.     

A high-resolution genetic map around waltzer on mouse Chromosome 10 and identification of a new allele of waltzer

A new autosomal recessive mouse mutation characterized by deafness and circling behavior was recovered during mutagenesis experiments with chlorambucil (CHL). On the basis of allelism tests and linkage analyses, this mutation appears to represent a new allele of waltzer (v) that maps to mouse Chromosome (Chr) 10. We have designated this new allele, Albany waltzer (v ^Alb ). A high-resolution map of the region around v was constructed from data from two intersubspecific backcrosses involving Mus musculus castaneus. The analysis of 648 backcross mice has allowed v ^Alb to be localized 1.1 ± 0.4 cM distal to D10Mit60 and 0.2 ± 0.2 cM proximal to a cluster of four markers, D10Mit172, D10Mit112, D10Mit48, and D10Mit196. An independent backcross was used to confirm the map order and distances in the v ^Alb backcross. The two linkage maps were consistent, indicating that the lesion in v ^Alb , which is presumed to be a deletion based on the known action of CHL, is small and has not significantly altered the map at this level of detection. Additionally, three genes (Ros1, Grik2, and Zfa) were eliminated as possible candidates for v ^Alb , and several SSLP markers were separated genetically. Received: 3 July 1996 / Accepted: 13 August 1996     

Location ofMls locus on mouse chromosome 1

Linkage between theMls locus and the chromosome 1 markersDip-1 andald was detected using two sets of recombinant inbred strains. Linkage betweenMls andDip-1 was confirmed in the fifth and sixth backcross generations of an incipient congenic strain. The AKXL data indicate that the gene order isDip-1-ald-Mls. The recombination frequency betweenald andMls is estimated to be 0.07 ±0.05, based on the AKXL data. The recombination frequency betweenDip-1 andMls is estimated to be 0.18 ±0.04, based on all the available data.     

Gypsy homologous sequences inDrosophila subobscura (gypsyDS)

Summary Characterization of sequences homologous to theDrosophila melanogaster gypsy transposable element was carried out inDrosophila subobscura (gypsyDS). They were found to be widely distributed among natural populations of this species. From Southern blot and in situ analyses, these sequences appear to be mobile in this species.GypsyDS sequences are located in both euchromatic and heterochromatic regions. A completegypsyDS sequence was isolated from aD. subobscura genomic library, and a 1.3-kb fragment which aligns with the ORF2 of theD. melanogaster gypsy element was sequenced. Comparisons of this sequence in three species (D. subobscura, D. melanogaster, and D. virilis) indicate that there is greater similarity between theD. subobscura-D. virilis sequences than betweenD. subobscura andD. melanogaster. Molecular divergence ofgypsy sequences betweenD. virilis andD. subobscura is estimated at 16 MY, whereas the most likely divergence time of these two species is more than 60 MY. These data strongly suggest thatgypsy sequences have been horizontally transferred between these species.Offprint requests to: T.M. Alberola     

Mapping of theHox-3.1 andMyc-1.2 genes on chromosome 15 of the mouse by restriction fragment length variations

Restriction endonuclease fragment length variations (RFLV) were detected by use of the cDNA probeHox-3.1 for the homeo box-3.1 gene and also thec-myc oncogene probe for exon 2. RFLV ofHox-3.1 were found inHindIII restriction patterns, and RFLV of theMyc-1.2 gene inEcoRV patterns. From the RFLV, theHox-3.1 andMyc-1.2 genes were mapped on chromosome 15. Three-point cross test data showed that the frequency of recombination is 26.4% betweenMyc-1.2 andGpt-1, 30.2% betweenGpt-1 andGdc-1, and 9.4% betweenGdc-1 andHox-3.1. The following order of these genes is proposed,Myc-1.2—Gpt-1—Gdc-1—Hox-3.1. All laboratory strains carry theHox-3.1 ^a andMyc-1.2 ^a alleles. Among strains of wild origin,domesticus strains carry only theHox-3.1 ^a andMyc-1.2 ^a alleles, as do the laboratory strains. One strain ofbrevirostris carries theHox-3.1 ^a andMyc-1.2 ^b alleles. Other wild subspecies from Europe and Asia,M. m. musculus, M. m. castaneus, M. m. molossinus, Chinese mice of wild origin, andM. m. yamashinai carry theHox-3.1 ^b andMyc-1.2 ^b alleles.     

Construction of a high-resolution genetic map encompassing the hotfoot locus

Hotfoot (ho) is a mutation affecting posture and movement. We report a new allele associated with the insertion of a transgene and its high-resolution mapping. Analysis of the transgene revealed that two complete and two truncated copies are inserted at the ho locus. The ho locus cosegregated with D6Mit299 in 702 meioses and is confined to a 1.1-cM region between the markers D6Mit122 and D6Mit174. If the order and distances between markers are consistent with previously published mapping data, the position of the ho locus must be revised and placed approximately 30 cM from the centromere. This high-resolution genetic map is the first step towards the positional cloning of the ho mutation. Received: 20 January 1997 / Accepted: 23 July 1997     

The ter primordial germ cell deficiency mutation maps near Grl-1 on mouse Chromosome 18

A single recessive gene, ter (teratoma), causes germ cell deficiency and a high incidence of congenital testicular teratomas in the 129/Sv-ter strain of the mouse. Linkage analyses between the ter gene and 36 marker genes of 19 chromosomes were performed with matings between the C57BL/6J-ter congenic strain and four inbred strains. Results showed that the ter gene was linked to D18Mit9, D18Mit14, and D18Mit17 on Chromosome (Chr) 18. Gene order estimated on the basis of recombination distance (in centimorgans) was [centromere-D18Mit14-5.1 (cM)-ter-0 (cM)-D18Mit17-23.8 (cM)-D18Mit9]. D18Mit17 is the microsatellite DNA of the Grl-1 (glucocorticoid receptor-1) locus. We conclude that the ter gene is closely linked to Grl-1 on Chr 18 and is a new mutation involving the developmental modification of primordial germ cells in mice.Deceased     

A High-Resolution Map in the Chromosomal Region Surrounding theLpsLocus

TheLpslocus on mouse chromosome 4 controls host responsiveness to lipopolysaccharide, a major component of the outer membrane of Gram-negative bacteria. The C3H/HeJ inbred mouse strain is characterized by a mutantLpsallele (Lps^d) that renders it hyporesponsive to LPS and naturally tolerant of its lethal effects. To identify theLpsgene by a positional cloning strategy, we have generated a high-resolution linkage map of the chromosomal region surrounding this locus. We have analyzed a total of 1604 backcross mice from a preexisting interspecific backcross panel of 259 (Mus spretus× C57BL/6J)F1 × C57BL/6J and two novel panels of 597 (DBA/2J × C3H/HeJ)F1 × C3H/HeJ and 748 (C57BL/6J × C3H/HeJ)F1 × C3H/HeJ segregating atLps.A total of 50 DNA markers have been mapped in a 11.8-cM span overlapping theLpslocus. This positions theLpslocus within a 1.1-cM interval, flanked proximally by a large cluster of markers, including three known genes (Cd30l, Hxb,andAmbp), and distally by two microsatellite markers (D4Mit7/D4Mit178). The localization of theLpslocus is several centimorgans proximal to that previously assigned.     

Localization of the bronx waltzer (bv) deafness gene to mouse Chromosome 5

The bronx waltzer (bv) mutation is an autosomal recessive mutation that is manifested as head tossing and circling in the mouse. The mutation affects the inner hair cells (IHCs) and pillar cells in the organ of Corti of the cochlea and the maculae and cristae of the vestibular part of the inner ear. IHCs begin to degenerate by a controlled mechanism of cell death as early as gestational day 17 (G17) in the basal coil of the cochlea, and few surviving IHCs are seen in the adult. As a first step towards the identification of bv, we analyzed a total of 20 loci in 118 mice from an intraspecific backcross giving the gene order: centromere–D5Mit1–D5Mit73–D5Mit55–[D5Mit12, Nds4 (Afp)]–D5Mit87–[D5Mit205, 20, 88, 208, 93–D5Mit338]–D5Mit25–D5Mit209–bv–D5Mit188–D5Mit367–D5Mit95–D5Mit43–D5Mit102. A total of 701 mice were then analyzed for the markers D5Mit93 and D5Mit95, defining a region of 12.08 cM flanking bv. Mice that were recombinant between D5Mit93 and D5Mit95 were analyzed for D5Mit338, D5Mit25, D5Mit209, bv, D5Mit188, and D5Mit367. bv maps 0.14 cM distal of the marker D5Mit209 and 1.14 cM proximal of the marker D5Mit188 in 701 backcross progeny. Received: 3 March 1997 / Accepted: 30 May 1997     

Rbt (rabo torcido), a new mouse skeletal mutation involved in anteroposterior patterning of the axial skeleton,maps close to the ts (tail-short) locus and distal to the sox9 locus on chromosome 11

Rbt (Rabo torcido) is a new semidominant mouse mutant with a variety of skeletal abnormalities. Heterozygous Rbt mutants display homeotic anteroposterior patterning problems along the axial skeleton that resemble Polycomb group and trithorax gene mutations. In addition, the Rbt mutant displays strong similarities to the phenotype observed in Ts (Tail-short), indicating also a homeotically transformed phenotype in these mice. We have mapped the Rbt locus to an interval of approximately 6 cM on mouse Chromosome (Chr) 11 between microsatellite markers D11Mit128 and D11Mit103. The Ts locus was mapped within a shorter interval of approximately 3 cM between D11Mit128 and D11Mit203. This indicates that Rbt and Ts may be allelic mutations. Sox9, the human homolog of which is responsible for the skeletal malformation syndrome campomelic dysplasia, was mapped proximal to D11Mit128. It is, therefore, unlikely that Ts and Rbt are mouse models for this human skeletal disorder. Received: 14 April 1996 / Accepted: 22 July 1996     

Localization of the mouse kidney disease (kd) gene to a YAC/BAC contig on Chromosome 10

Mice that are homozygous for the kidney disease (kd) gene on Chromosome (Chr) 10 spontaneously develop a progressive and fatal interstitial nephritis. The disease phenotype is similar to that of the human disease, juvenile nephronophthisis. Using a backcross and intercross breeding strategy and analysis of over 900 resultant progeny, this genetic locus has now been mapped to a minimal co-segregating region of approximately two megabases between D10Mit 193 and D10Mit 38. The location assigned to kd by this study is over 3 cM from the current Mouse Genome Database location. The entire interval has been cloned in yeast artificial chromosome (YAC) and bacterial artificial chromosome (BAC) clones. Recombinant analysis has permitted assignment of 13 Mit microsatellite markers to positions near or within the region. Two new markers have been identified by using single-strand conformation polymorphism (SSCP) analysis of sequenced BAC ends. Several BAC end sequences align with human BAC clones from Chr 6q21 that contain NR2E1, Snx3, and Ros1. Three murine genes, CD24a, fyn, and ColX reported to map in or near the kd region as defined by this study have been evaluated. Though not definitely excluded, they appear to be unlikely candidates. Received: 23 July 1999 / Accepted 23 June 2000     

Mapping of the Mouse Actin Capping Protein α Subunit Genes and Pseudogenes

Capping protein (CP), a heterodimer of α and β subunits, is found in all eukaryotes. CP binds to the barbed ends of actin filamentsin vitroand controls actin assembly and cell motilityin vivo.Vertebrates have three α isoforms (α1, α2, α3) produced from different genes, whereas lower organisms have only one gene and one isoform. We isolated genomic clones corresponding to the α subunits of mouse CP and found three α1 genes, two of which are pseudogenes, and a single gene for both α2 and α3. Their chromosomal locations were identified by interspecies backcross mapping. The α1 gene (Cappa1) mapped to Chromosome 3 betweenD3Mit11andD3Mit13.The α1 pseudogenes (Cappa1-ps1andCappa1-ps2) mapped to Chromosomes 1 and 9, respectively. The α2 gene (Cappa2) mapped to Chromosome 6 nearPtn.The α3 gene (Cappa3) also mapped to Chromosome 6, approximately 68 cM distal fromCappa2nearKras2.One mouse mutation,de,maps in the vicinity of the α1 gene. No known mouse mutations map to regions near the α2 or α3 genes.