Identification of Mom7, a novel modifier of Apc(Min/+) on mouse chromosome 18

The Apc(Min) mouse model of colorectal cancer provides a discrete, quantitative measurement of tumor multiplicity, allowing for robust quantitative trait locus analysis. This advantage has previously been used to uncover polymorphic modifiers of the Min phenotype: Mom1, which is partly explained by Pla2g2a; Mom2, a spontaneous mutant modifier; and Mom3, which was discovered in an outbred cross. Here, we describe the localization of a novel modifier, Mom7, to the pericentromeric region of chromosome 18. Mom7 was mapped in crosses involving four inbred strains: C57BL/6J (B6), BTBR/Pas (BTBR), AKR/J (AKR), and A/J. There are at least two distinct alleles of Mom7: the recessive, enhancing BTBR, AKR, and A/J alleles and the dominant, suppressive B6 allele. Homozygosity for the enhancing alleles increases tumor number by approximately threefold in the small intestine on both inbred and F(1) backgrounds. Congenic line analysis has narrowed the Mom7 region to within 7.4 Mb of the centromere, 28 Mb proximal to Apc. Analysis of SNP data from various genotyping projects suggests that the region could be as small as 4.4 Mb and that there may be five or more alleles of Mom7 segregating among the many strains of inbred mice. This has implications for experiments involving Apc(Min) and comparisons between different or mixed genetic backgrounds.     

Hippocampal commissure defects in crosses of four inbred mouse strains with absent corpus callosum

It is known that four common inbred mouse strains show defects of the forebrain commissures. The BALB/cJ strain has a low frequency of abnormally small corpus callosum, whereas the 129 strains have many animals with deficient corpus callosum. The I/LnJ and BTBR T+ tf/J strains never have a corpus callosum, whereas half of I/LnJ and almost all BTBR show severely reduced size of the hippocampal commissure. Certain F1 hybrid crosses among these strains are known to be less severely abnormal than the inbred parents, suggesting that the parent strains have different genetic causes of commissure defects. In this study, all hybrid crosses among the four strains were investigated. The BTBR × I/Ln hybrid expressed almost no defects of the hippocampal commissure, unlike its inbred parent strains. Numerous three‐way crosses among the four strains yielded many mice with no corpus callosum and severely reduced hippocampal commissure, which shows that the phenotypic defect can result from several different combinations of genetic alleles. The F2 and F3 hybrid crosses of BTBR and I/LnJ had almost 100% absence of the corpus callosum but about 50% frequency of deficient hippocampal commissure. The four‐way hybrid cross among all four abnormal strains involved highly fertile parents and yielded a very wide phenotypic range of defects from almost no hippocampal commissure to totally normal forebrain commissures. The F2 and F3 crosses as well as the four‐way cross provide excellent material for studies of genetic linkage and behavioral consequences of commissure defects.     

Recombinant inbreeding in mice reveals thresholds in embryonic corpus callosum development

The inbred strains BALB/cWah1 and 129P1/ReJ both show incomplete penetrance for absent corpus callosum (CC); about 14% of adult mice have no CC at all. Their F(1) hybrid offspring are normal, which proves that the strains differ at two or more loci pertinent to absent CC. Twenty-three recombinant inbred lines were bred from the F(2) cross of BALB/c and 129, and several of these expressed a novel and severe phenotype after only three or four generations of inbreeding - total absence of the CC and severe reduction of the hippocampal commissure (HC) in every adult animal. As inbreeding progressed, intermediate sizes of the CC and the HC remained quite rare. This striking phenotypic distribution in adults arose from developmental thresholds in the embryo. CC axons normally cross to the opposite hemisphere via a tissue bridge in the septal region at midline, where the HC forms before CC axons arrive. The primary defect in callosal agenesis in the BALB/c and 129 strains is severe retardation of fusion of the hemispheres in the septal region, and failure to form a CC is secondary to this defect. The putative CC axons arrive at midline at the correct time and place in all groups, but in certain genotypes, the bridge is not yet present. The relative timing of axon growth and delay of the septal bridge create a narrow critical period for forming a normal brain.     

A genetic linkage map of Cryptococcus neoformans variety neoformans serotype D (Filobasidiella neoformans)

To construct a genetic linkage map of the heterothallic yeast, Cryptococcus neoformans (Filobasidiella neoformans), we crossed two mating-compatible strains and analyzed 94 progeny for the segregation of 301 polymorphic markers, consisting of 228 restriction site polymorphisms, 63 microsatellites, two indels, and eight mating-type (MAT)-associated markers. All but six markers showed no significant (P < 0.05) segregation distortion. At a minimum LOD score of 6.0 and a maximum recombination frequency of 0.30, 20 linkage groups were resolved, resulting in a map length of approximately 1500 cM. Average marker density is 5.4 cM (range 1-28.7 cM). Hybridization of selected markers to blots of electrophoretic karyotypes unambiguously assigned all linkage groups to chromosomes and led us to conclude that the C. neoformans genome is approximately 20.2 Mb, comprising 14 chromosomes ranging in size from 0.8 to 2.3 Mb, with a ratio of approximately 13.2 kb/cM averaged across the genome. However, only 2 of 12 ungrouped markers hybridized to chromosome 10. The hybridizations revealed at least one possible reciprocal translocation involving chromosomes 8, 9, and 12. This map has been critical to genome sequence assembly and will be essential for future studies of quantitative trait inheritance.     

Positional cloning of a type 2 diabetes quantitative trait locus; tomosyn-2, a negative regulator of insulin secretion

We previously mapped a type 2 diabetes (T2D) locus on chromosome 16 (Chr 16) in an F2 intercross from the BTBR T (+) tf (BTBR) Lep(ob/ob) and C57BL/6 (B6) Lep(ob/ob) mouse strains. Introgression of BTBR Chr 16 into B6 mice resulted in a consomic mouse with reduced fasting plasma insulin and elevated glucose levels. We derived a panel of sub-congenic mice and narrowed the diabetes susceptibility locus to a 1.6 Mb region. Introgression of this 1.6 Mb fragment of the BTBR Chr 16 into lean B6 mice (B6.16(BT36-38)) replicated the phenotypes of the consomic mice. Pancreatic islets from the B6.16(BT36-38) mice were defective in the second phase of the insulin secretion, suggesting that the 1.6 Mb region encodes a regulator of insulin secretion. Within this region, syntaxin-binding protein 5-like (Stxbp5l) or tomosyn-2 was the only gene with an expression difference and a non-synonymous coding single nucleotide polymorphism (SNP) between the B6 and BTBR alleles. Overexpression of the b-tomosyn-2 isoform in the pancreatic β-cell line, INS1 (832/13), resulted in an inhibition of insulin secretion in response to 3 mM 8-bromo cAMP at 7 mM glucose. In vitro binding experiments showed that tomosyn-2 binds recombinant syntaxin-1A and syntaxin-4, key proteins that are involved in insulin secretion via formation of the SNARE complex. The B6 form of tomosyn-2 is more susceptible to proteasomal degradation than the BTBR form, establishing a functional role for the coding SNP in tomosyn-2. We conclude that tomosyn-2 is the major gene responsible for the T2D Chr 16 quantitative trait locus (QTL) we mapped in our mouse cross. Our findings suggest that tomosyn-2 is a key negative regulator of insulin secretion.     

A high‐density linkage map enables a second‐generation collared flycatcher genome assembly and reveals the patterns of avian recombination rate variation and chromosomal evolution

Detailed linkage and recombination rate maps are necessary to use the full potential of genome sequencing and population genomic analyses. We used a custom collared flycatcher 50 K SNP array to develop a high‐density linkage map with 37 262 markers assigned to 34 linkage groups in 33 autosomes and the Z chromosome. The best‐order map contained 4215 markers, with a total distance of 3132 cM and a mean genetic distance between markers of 0.12 cM . Facilitated by the array being designed to include markers from most scaffolds, we obtained a second‐generation assembly of the flycatcher genome that approaches full chromosome sequences (N50 super‐scaffold size 20.2 Mb and with 1.042 Gb (of 1.116 Gb) anchored to and mostly ordered and oriented along chromosomes). We found that flycatcher and zebra finch chromosomes are entirely syntenic but that inversions at mean rates of 1.5–2.0 event (6.6–7.5 Mb) per My have changed the organization within chromosomes, rates high enough for inversions to potentially have been involved with many speciation events during avian evolution. The mean recombination rate was 3.1 cM /Mb and correlated closely with chromosome size, from 2 cM /Mb for chromosomes >100 Mb to >10 cM /Mb for chromosomes <10 Mb. This size dependence seemed entirely due to an obligate recombination event per chromosome; if 50 cM was subtracted from the genetic lengths of chromosomes, the rate per physical unit DNA was constant across chromosomes. Flycatcher recombination rate showed similar variation along chromosomes as chicken but lacked the large interior recombination deserts characteristic of zebra finch chromosomes.     

Variation in the ratio of physical to genetic distance in intervals adjacent to theMla locus on barley chromosome 1H

Variants of the pulsed-field gel electrophoresis technique were used in conjunction with two-dimensional DNA gel electrophoresis (2-DDGE) to determine the ratio of physical to genetic distance in two genetically defined intervals on barley chromosome 1H.2-DDGE analysis demonstrated that two loci that define a 0.3 cM interval, as determined by hybridization with BCD249, reside on a single 450-kbMluI fragment. This result indicates a maximum ratio of physical to genetic distance in this interval of 1500 kb/cM as compared to 3.7–4.2 Mb/cM for the barley genome as a whole. High molecular weight (HMW) DNA restricted withNotI and probed sequentially with MWG068 and BCD249 yield diffuse bands at approximately 2.8 Mb and 3.0 Mb in the C.I. 16151 and C.I. 16155 parental lines, respectively. These results suggest the maximum ratio of physical to genetic distance in the interval defined by these probes is 7.8 Mb/cM. unique HMW DNA restriction fragment length polymorphisms (RFLP) were attributed to the presence of recombination breakpoints. Data from the recombination breakpoint analysis were used to estimate a ratio of physical to genetic distance of 2.5 Mb/cM in theXbcd249.2-Xmwg068 interval and 0.465 Mb/cM in theXbcd249.1-Xbcd249.2 interval. Both physical linkage and recombination breakpoint analysis indicate theXbcd249.1-Xbcd249.2 interval is approximately five-fold smaller, physically, than theXbcd249.2-Xmwg068 interval.     

X chromosome influences sperm length in the stalk-eyed fly Cyrtodiopsis dalmanni

Whether sexually selected traits are sex linked can have profound effects on their evolution. In the diopsid stalk-eyed fly, Cyrtodiopsis dalmanni, sperm length and female reproductive tract morphology have coevolved across species, postcopulatory sexual selection is known to occur, and X-linked genes affect female ventral sperm receptacle size. Here, we estimate the location of quantitative trait loci (QTL) for spermatocyst tail length by using F2 progeny segregating for an X-linked factor that causes sex-ratio meiotic drive. We found two QTL, including a major X-linked QTL responsible for 25% of the variation in spermatocyst tail length 2.1 cM from the sex-ratio element and 0.8 cM from a major eye span QTL. Sex-ratio males produce shorter spermatocyst tails and shorter eye spans. Thus, X-linked factors affect both pre- and postcopulatory traits, and linkage between the alleles for short eye span and short spermatocyst tail length allow pre- and postcopulatory sexual selection to potentially act in concert against the transmission bias caused by the sex-ratio chromosome.     

Variation in the ratio of physical to genetic distance in intervals adjacent to theMla locus on barley chromosome 1H

Variants of the pulsed-field gel electrophoresis technique were used in conjunction with two-dimensional DNA gel electrophoresis (2-DDGE) to determine the ratio of physical to genetic distance in two genetically defined intervals on barley chromosome 1H.2-DDGE analysis demonstrated that two loci that define a 0.3 cM interval, as determined by hybridization with BCD249, reside on a single 450-kbMluI fragment. This result indicates a maximum ratio of physical to genetic distance in this interval of 1500 kb/cM as compared to 3.7–4.2 Mb/cM for the barley genome as a whole. High molecular weight (HMW) DNA restricted withNotI and probed sequentially with MWG068 and BCD249 yield diffuse bands at approximately 2.8 Mb and 3.0 Mb in the C.I. 16151 and C.I. 16155 parental lines, respectively. These results suggest the maximum ratio of physical to genetic distance in the interval defined by these probes is 7.8 Mb/cM. unique HMW DNA restriction fragment length polymorphisms (RFLP) were attributed to the presence of recombination breakpoints. Data from the recombination breakpoint analysis were used to estimate a ratio of physical to genetic distance of 2.5 Mb/cM in theXbcd249.2-Xmwg068 interval and 0.465 Mb/cM in theXbcd249.1-Xbcd249.2 interval. Both physical linkage and recombination breakpoint analysis indicate theXbcd249.1-Xbcd249.2 interval is approximately five-fold smaller, physically, than theXbcd249.2-Xmwg068 interval.Names are necessary to report factually on available data; however the USDA neither guarantees nor warrants the standard of the product and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable     

A Restriction Fragment Length Polymorphism Map and Electrophoretic Karyotype of the Fungal Maize Pathogen Cochliobolus Heterostrophus

A restriction fragment length polymorphism (RFLP) map has been constructed of the nuclear genome of the plant pathogenic ascomycete Cochliobolus heterostrophus. The segregation of 128 RFLP and 4 phenotypic markers was analyzed among 91 random progeny of a single cross; linkages were detected among 126 of the markers. The intact chromosomal DNAs of the parents and certain progeny were separated using pulsed field gel electrophoresis and hybridized with probes used to detect the RFLPs. In this way, 125 markers were assigned to specific chromosomes and linkages among 120 of the markers were confirmed. These linkages totalled 941 centimorgans (cM). Several RFLPs and a reciprocal translocation were identified tightly linked to Tox1, a locus controlling host-specific virulence. Other differences in chromosome arrangement between the parents were also detected. Fourteen gaps of at least 40 cM were identified between linkage groups on the same chromosomes; the total map length was therefore estimated to be, at a minimum, 1501 cM. Fifteen A chromosomes ranging from about 1.3 megabases (Mb) to about 3.7 Mb were identified; one of the strains also has an apparent B chromosome. This chromosome appears to be completely dispensable; in some progeny, all of 15 markers that mapped to this chromosome were absent. The total genome size was estimated to be roughly 35 Mb. Based on these estimates of map length and physical genome size, the average kb/cM ratio in this cross was calculated to be approximately 23. This low ratio of physical length to map distance should make this RFLP map a useful tool for cloning genes.     

Identification and sequencing the juvenile spermatogonial depletion critical interval on mouse chromosome 1 reveals the presence of eight candidate genes.

In mice, the recessive, non-pleiotropic, juvenile spermatogonial depletion (jsd) mutation results in a single wave of spermatogenesis, followed by failure of type A spermatogonial stem cells to differentiate, rendering adult males sterile. As part of an effort to identify the gene underlying this mutation, we report here the construction of a high-resolution genetic map involving more than 1000 meioses and 24 polymorphic loci. Our data define a critical jsd interval of approximately 0.4 cM at 49 cM on mouse chromosome 1, between D1Mit215 and 257SP6. We have constructed a physical map spanning the region comprising 24 overlapping BACs. Eighteen of these BACs have been fully sequenced, or are in draft form, allowing us to annotate approximately 2.5 Mb of DNA surrounding the jsd locus. The critical 0.4 cM jsd interval corresponds to a physical distance of approximately 1.5 Mb. Eight genes have been identified in this interval, two of which appear to be possible candidates for the jsd mutation.     

Aberrant immune responses in a mouse with behavioral disorders

BTBR T+tf/J (BTBR) mice have recently been reported to have behaviors that resemble those of autistic individuals, in that this strain has impairments in social interactions and a restricted repetitive and stereotyped pattern of behaviors. Since immune responses, including autoimmune responses, are known to affect behavior, and individuals with autism have aberrant immune activities, we evaluated the immune system of BTBR mice, and compared their immunity and degree of neuroinflammation with that of C57BL/6 (B6) mice, a highly social control strain, and with F1 offspring. Mice were assessed at postnatal day (pnd) 21 and after behavioral analysis at pnd70. BTBR mice had significantly higher amounts of serum IgG and IgE, of IgG anti-brain antibodies (Abs), and of IgG and IgE deposited in the brain, elevated expression of cytokines, especially IL-33 IL-18, and IL-1β in the brain, and an increased proportion of MHC class II-expressing microglia compared to B6 mice. The F1 mice had intermediate levels of Abs and cytokines as well as social activity. The high Ab levels of BTBR mice are in agreement with their increased numbers of CD40(hi)/I-A(hi) B cells and IgG-secreting B cells. Upon immunization with KLH, the BTBR mice produced 2-3 times more anti-KLH Abs than B6 mice. In contrast to humoral immunity, BTBR mice are significantly more susceptible to listeriosis than B6 or BALB/c mice. The Th2-like immune profile of the BTBR mice and their constitutive neuroinflammation suggests that an autoimmune profile is implicated in their aberrant behaviors, as has been suggested for some humans with autism.     

How strong is the mutagenicity of recombination in mammals?

It is commonly believed that a high recombination rate such as that in a pseudoautosomal region (PAR) greatly increases the mutation rate because a 170-fold increase was estimated for the mouse PAR region. However, sequencing PAR and non-PAR introns of the Fxy gene in four Mus taxa, we found an increase of only twofold to fivefold. Furthermore, analyses of sequence data from human and orangutan PAR and X-linked regions and from autosomal regions showed a weak effect of recombination on mutation rate (a slope of less than 0.2% per cM/Mb), although a much stronger effect on GC content (1% to 2% per cM/Mb). Because typical recombination rates in mammals are much lower than those in PARs, the mutagenicity of recombination is weak or, at best, moderate, although its effect on GC% is much stronger. In addition, contrary to a previous study, we found no Fxy duplicate in Mus spretus.     

Physical Mapping of Rice Chromosomes 4 and 7 Using YAC Clones

Physical maps of rice chromosomes 4 and 7 were constructed bylanding yeast artificial chromosomes (YACs) along our high-densitymolecular linkage map. Using 114 DNA markers, 258 individualYACs were located on chromosome 4. Sixty-two out of 258 YACscarried two or more DNA marker positions and formed 16 contigswhich covered a total length of 17.1 cM. The other YACs werearranged to 23 positions. On chromosome 7, 203 individual YACswere landed on 109 DNA markers. Sixty-four out of 203 YACs formed15 contigs which covered a total length of 21.8 cM and 139 YACswere localized to 26 positions. Chromosomes 4 and 7 were coveredwith minimum tiling paths of 45 and 48 YACs, respectively. Takingthe average size of YAC insert DNA to be 350 kb and the entiregenome size to be 430 Mb, about 16–18 Mb of each chromosomeor an estimated 50% of their total lengths have been coveredwith YACs. Physical maps of these 2 chromosomes should be ofgreat help in identifying useful trait genes and unravelinggenetic and biological characteristics in rice.     

A member of a gene family on Xp22.3, VCX-A, is deleted in patients with X-linked nonspecific mental retardation

X-linked nonspecific mental retardation (MRX) has a frequency of 0.15% in the male population and is caused by defects in several different genes on the human X chromosome. Genotype-phenotype correlations in male patients with a partial nullisomy of the X chromosome have suggested that at least one locus involved in MRX is on Xp22.3. Previous deletion mapping has shown that this gene resides between markers DXS1060 and DXS1139, a region encompassing approximately 1.5 Mb of DNA. Analyzing the DNA of 15 males with Xp deletions, we were able to narrow this MRX critical interval to approximately 15 kb of DNA. Only one gene, VCX-A (variably charged, X chromosome mRNA on CRI-S232A), was shown to reside in this interval. Because of a variable number of tandem 30-bp repeats in the VCX-A gene, the size of the predicted protein is 186-226 amino acids. VCX-A belongs to a gene family containing at least four nearly identical paralogues on Xp22.3 (VCX-A, -B, -B1, and -C) and two on Yq11.2 (VCY-D, VCY-E), suggesting that the X and Y copies were created by duplication events. We have found that VCX-A is retained in all patients with normal intelligence and is deleted in all patients with mental retardation. There is no correlation between the presence or absence of VCX-B1, -B, and VCX-C and mental status in our patients. These results suggest that VCX-A is sufficient to maintain normal mental development.     

Genomewide linkage scan for split-hand/foot malformation with long-bone deficiency in a large Arab family identifies two novel susceptibility loci on chromosomes 1q42.2-q43 and 6q14.1

Split-hand/foot malformation with long-bone deficiency (SHFLD) is a rare, severe limb deformity characterized by tibia aplasia with or without split-hand/split-foot deformity. Identification of genetic susceptibility loci for SHFLD has been unsuccessful because of its rare incidence, variable phenotypic expression and associated anomalies, and uncertain inheritance pattern. SHFLD is usually inherited as an autosomal dominant trait with reduced penetrance, although recessive inheritance has also been postulated. We conducted a genomewide linkage analysis, using a 10K SNP array in a large consanguineous family (UR078) from the United Arab Emirates (UAE) who had disease transmission consistent with an autosomal dominant inheritance pattern. The study identified two novel SHFLD susceptibility loci at 1q42.2-q43 (nonparametric linkage [NPL] 9.8, P=.000065) and 6q14.1 (NPL 7.12, P=.000897). These results were also supported by multipoint parametric linkage analysis. Maximum multipoint LOD scores of 3.20 and 3.78 were detected for genomic locations 1q42.2-43 and 6q14.1, respectively, with the use of an autosomal dominant mode of inheritance with reduced penetrance. Haplotype analysis with informative crossovers enabled mapping of the SHFLD loci to a region of approximately 18.38 cM (8.4 Mb) between single-nucleotide polymorphisms rs1124110 and rs535043 on 1q42.2-q43 and to a region of approximately 1.96 cM (4.1 Mb) between rs623155 and rs1547251 on 6q14.1. The study identified two novel loci for the SHFLD phenotype in this UAE family.     

An integrated map of human 6q22.3-q24 including a 3-Mb high-resolution BAC/PAC contig encompassing a QTL for fetal hemoglobin

Genetic studies have previously assigned a quantitative trait locus (QTL) for hemoglobin F and F cells to a region of approximately 4 Mb between the markers D6S408 and D6S292 on chromosome 6q23. An initial yeast artificial chromosome contig of 13 clones spanning this region was generated. Further linkage analysis of an extended kindred refined the candidate interval to 1-2 cM, and key recombination events now place the QTL within a region of <800 kb. We describe a high-resolution bacterial clone contig spanning 3 Mb covering this critical region. The map consists of 223 bacterial artificial chromosome (BAC) and 100 P1 artificial chromosome (PAC) clones ordered by sequence-tagged site (STS) content and restriction fragment fingerprinting with a minimum tiling path of 22 BACs and 1 PAC. A total of 194 STSs map to this interval of 3 Mb, giving an average marker resolution of approximately one per 15 kb. About half of the markers were novel and were isolated in the present study, including three CA repeats and 13 single nucleotide polymorphisms. Altogether 24 expressed sequence tags, 6 of which are unique genes, have been mapped to the contig.     

Estimation of genetic distances between “reeler” and nearby loci on mouse chromosome 5

Genetic distances between reeler (gene symbol rl) and the adjacent loci Pgy-1, Sor, and En-2 on proximal chromosome 5 were estimated using a backcross panel between rlOrl/rlOrl BALB/c and C57 mice. Pgy-1 and Sor are located approximately 7 cM away from and centromeric to reeler, whereas En-2 is located distally, approximately 8 cM from reeler. Backcrosses between rlOrl/rlOrl BALB/c and mice with the T31H translocation showed that the breakpoint is located less than 2 cM from reeler. Together with previous work, these observations suggest the following gene order: Cen-Sor/Pgy-1-rl-T31H-En-2-other loci.     

A bacterial artificial chromosome library for 'Jefferson' hazelnut and identification of clones associated with eastern filbert blight resistance and pollen-stigma incompatibility

European hazelnut (Corylus avellana L.) is the only economically important nut crop in the family Betulaceae. Because of its small genome size (~385 Mb / 1C), relatively short life cycle, availability of a dense linkage map, and amenability to transformation by Agrobacterium, the European hazelnut could serve as a model plant for the Betulaceae. Here we report the construction of a bacterial artificial chromosome (BAC) library for 'Jefferson' hazelnut using the cloning enzyme MboI and the vector pECBAC1 (BamHI site). The library consists of 39,936 clones arrayed in 104,384-well microtitre plates with a mean insert size of 117 kb. The genomic coverage of the library is estimated to be about 12 genome equivalents. This library provides a valuable resource for the map-based cloning of two important genes, the resistance gene from 'Gasaway' that confers resistance to eastern filbert blight caused by the fungus Anisogramma anomala (Peck) E. Müller and the S locus that controls pollen-stigma incompatibility. Fine-resolution mapping near the two loci was carried out using random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers. Fine mapping at the disease resistance locus showed that markers W07-375 and X01-825 flanked the resistance locus at distances of 0.06 and 0.05 cM, respectively. The S locus is flanked by markers 204-950 and KG819-200 at distances of 0.14 and 0.24 cM, respectively. Assuming that 1 cM corresponds to a physical distance of 430 kb, it will take approximately two to three chromosome walks to assemble BAC contigs that span both loci.     

Linkage analysis of X-linked cone-rod dystrophy: localization to Xp11.4 and definition of a locus distinct from RP2 and RP3.

Progressive X-linked cone-rod dystrophy (COD1) is a retinal disease affecting primarily the cone photoreceptors. The COD1 locus originally was localized, by the study of three independent families, to a region between Xp11.3 and Xp21.1, encompassing the retinitis pigmentosa (RP) 3 locus. We have refined the COD1 locus to a limited region of Xp11.4, using two families reported elsewhere and a new extended family. Genotype analysis was performed by use of eight microsatellite markers (tel-M6CA, DXS1068, DXS1058, DXS993, DXS228, DXS1201, DXS1003, and DXS1055-cent), spanning a distance of 20 cM. Nine-point linkage analysis, by use of the VITESSE program for X-linked disorders, established a maximum LOD score (17.5) between markers DXS1058 and DXS993, spanning 4.0 cM. Two additional markers, DXS977 and DXS556, which map between DXS1058 and DXS993, were used to further narrow the critical region. The RP3 gene, RPGR, was excluded on the basis of two obligate recombinants, observed in two independent families. In a third family, linkage analysis did not exclude the RPGR locus. The entire coding region of the RPGR gene from two affected males from family 2 was sequenced and was found to be normal. Haplotype analysis of two family branches, containing three obligate recombinants, two affected and one unaffected, defined the COD1 locus as distal to DXS993 and proximal to DXS556, a distance of approximately 1.0 Mb. This study excludes COD1 as an allelic variant of RP3 and establishes a novel locus that is sufficiently defined for positional cloning.