A 1.5-Mb physical map of the hidrotic ectodermal dysplasia (Clouston syndrome) gene region on human chromosome 13q11

The HED (hidrotic ectodermal dysplasia) or Clouston syndrome gene (named ED2) has been mapped to the pericentromeric region of chromosome 13 (13q11) to a 2.4-cM interval flanked by markers D13S1828 and D13S1830. We have developed a BAC/PAC-based contig map of this region. This contig, comprising 23 clones and spanning 1.5 Mb, was established by mapping of 27 BAC/PAC end-derived STSs, 11 known polymorphic markers, 2 previously mapped genes, and 14 ESTs. The genomic clone overlaps were confirmed by restriction fragment fingerprint analysis. This contig provides the basis for genomic sequencing and gene identification in the ED2 critical region. Of the 14 ESTs mapped to the contig, 6 show homology to human genes and 8 appear to be novel. Expression patterns of the genes/ESTs were tested by Northern blot and RT-PCR. Full characterization of some of these genes, as well as the novel ESTs, will be useful in assessing their involvement in the HED/Clouston syndrome.     

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS): high-resolution physical and transcript map of the candidate region in chromosome region 13q11

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS or SACS) is a neurodegenerative disease frequent in northeastern Québec. In a previous study, we localized the disease gene to chromosome region 13q11 by identifying excess sharing of a marker allele in patients followed by linkage analysis and haplotyping. To create a detailed physical map of this region, we screened CEPH mega-YACs with 41 chromosome 13 sequence-tagged-sites (STSs) known to map to 13q11-q12. The YAC contig, composed of 27 clones, extends on the genetic map from D13S175 to D13S221, an estimated distance of at least 19.3 cM. A high-resolution BAC and PAC map that includes the ARSACS critical region flanked by D13S1275 and D13S292 was constructed. These YAC and BAC/PAC maps allowed the accurate placement of 29 genes and ESTs previously mapped to the proximal region of chromosome 13q. We confirmed the position of two candidate genes within the critical region and mapped the other 27 genes and ESTs to nearby intervals. Six BAC/PAC clones form a contig between D13S232 and D13S787 for sequencing within the ARSACS critical region.     

Assignment of 48 ESTs to Chromosome 13 Band q14.3 and Expression Pattern for ESTs Located in the Core Region Deleted in B-CLL

An expression map containing 48 ESTs was constructed to identify a tumor-suppressor gene involved in B-cell chronic lymphocytic leukemia (B-CLL), which was previously assigned to chromosome band 13q14.3 close to genetic markers D13S25 and D13S319. Thirty-nine of these 48 ESTs, together with 11 additional ones listed in databases, were initially assigned to chromosome 13q14 between markers D13S168 and D13S176. Nine others have recently been located in the D13S319 region. Our results indicate that 48 of the 59 ESTs analyzed belong to a YAC contig of chromosome 13 band q14, and 22 are contained on YAC 933e9, which encompasses the B-CLL critical region. Ten of these 22 ESTs were accurately assigned on a PAC, BAC, and cosmid contig encompassing the smallest minimal deletion area described so far in B-CLL, and 20 were tested for their expression on 27 normal or tumor tissues. One EST appears to be a likely candidate for the tumor-suppressor gene involved in B-CLL.     

A physical map of the 20q12-q13.1 region associated with type 2 diabetes

Several recent genetic studies have suggested linkage of Type 2 diabetes (non-insulin-dependent diabetes mellitus) susceptibility to a region of chromosome 20q12-q13.1. To facilitate the identification and cloning of a diabetes susceptibility gene(s) in this region, we have constructed correlated radiation hybrid and YAC/BAC contig physical maps of the region. A high-resolution radiation hybrid map encompassing 9.5 Mb between the PLC and the CEBPB genes was constructed using 68 markers: 25 polymorphic markers, 15 known genes, 21 ESTs, and 7 random genomic sequences. The physical order of the polymorphic markers within this radiation hybrid map is consistent with published genetic maps. A YAC/BAC contig that gives continuous coverage between PLC and CEBPB was also constructed. This contig was constructed from 24 YACs, 34 BACs, and 1 P1 phage clone onto which 71 markers were mapped: 23 polymorphic markers, 12 genes, 24 ESTs, and 12 random genomic sequences. The radiation hybrid map and YAC/BAC physical map enable precise mapping of newly identified transcribed sequences and polymorphic markers that will aid in linkage and linkage disequilibrium studies and facilitate identification and cloning of candidate Type 2 diabetes susceptibility genes residing in 20q12-q13.1.     

A clone contig of 12q24.3 encompassing the distal hereditary motor neuropathy type II gene

We previously assigned the disease locus for autosomal dominant hereditary motor neuropathy type II (distal HMN II) within a 13-cM interval at chromosome 12q24.3. We constructed a physical map of the distal HMN II region based on yeast artificial chromosomes (YACs), P1 artificial chromosomes (PACs), and bacterial artificial chromosomes (BACs) using an STS content mapping approach. The contig contains 26 YAC, 15 PAC, and 60 BAC clones and covers a physical distance of approximately 5 Mb. A total of 99 STS markers, including 25 known STSs and STRs, 49 new STSs generated from clone end-fragments, 20 ESTs, and 5 known genes, were located on the contig. This physical map provides a valuable resource for mapping genes and markers located within the distal HMN II region and facilitates the positional cloning of the distal HMN II gene.     

Construction of a 1.2-Mb BAC/PAC contig of the porcine gene RYR1 region on SSC 6q1.2 and comparative analysis with HSA 19q13.13

We screened a porcine bacterial artificial chromosome (BAC) and a P1 derived artificial chromosome (PAC) library to construct a sequence-ready approximately 1.2-Mb BAC/PAC contig of the ryanodine receptor-1 gene (RYR1) region on porcine chromosome (SSC) 6q1.2. This genomic segment is of special interest because it harbors the locus for stress susceptibility in pigs and a putative quantitative trait locus for muscle growth. Detailed physical mapping of this gene-rich region allowed us to assign to this contig 17 porcine genes orthologous to known human chromosome 19 genes. Apart from the relatively well-characterized porcine gene RYR1, the other 16 genes represent novel chromosomal assignments and 14 genes have been cloned for the first time in pig. Comparative analysis of the porcine BAC/PAC contig with the human chromosome (HSA) 19q13.13 map revealed a completely conserved gene order of this segment between pig and human. A detailed porcine-human-mouse comparative map of this region was constructed.     

Construction of a physical and transcript map for a 1-Mb genomic region containing the urofacial (Ochoa) syndrome gene on 10q23-q24 and localization of the disease gene within two overlapping BAC clones (<360 kb).

Urofacial (Ochoa) syndrome is an autosomal recessive disease characterized by distorted facial expression and urinary abnormalities. Previously, we mapped the UFS gene to chromosome 10q23-q24 and narrowed the interval to one YAC clone of 1410 kb. Here, we have constructed a BAC/PAC contig of the 1-Mb region using STS content mapping with 42 BAC/PAC-end sequences, 9 previously reported and 16 newly identified microsatellite markers, and 14 EST markers. A total of 26 polymorphic microsatellite markers were genotyped for 31 UFS patients from Colombia and 2 patients from the United States. Haplotype analyses suggest that the UFS gene is located within two overlapping BAC clones, a region of <360 kb of DNA sequence. We tested 42 EST markers previously mapped to the D10S1709-D10S603 interval against the BAC/PAC contig and identified 11 ESTs located in the 1-Mb region. Four of the 11 ESTs mapped to the 360-kb UFS critical region. Shotgun sequencing of the two BAC clones and BLASTN search of the EST databases revealed 3 other ESTs contained in the UFS critical region. These results will facilitate the cloning and identification of the UFS gene.     

A transcript map of an 800-kb region on human chromosome 11q13, part of the candidate region for SCA5 and BBS1

The exon-amplification method was used to identify putative transcribed sequences from an 800-kb region that includes the genes for phospholipase Cβ3 and PYGM on human chromosome 11q13. The clone contig consisted of ten cosmids, three bacterial artificial chromosomes, and one P1 artificial chromosome. A total of 83 exons were generated of which 23 were derived from known genes and expressed sequence tags (ESTs). Five different EST cDNA clones were identified and mapped on the contig. One is a homolog of the human p70S6 kinase (p70s6 k) gene whose function involves the translational regulation of ribosomal protein synthesis and thereby impacts on ribosomal biogenesis. The gene for p70s6 k is expressed universally, including within adipose cells and retina, and it could play a role in Bardet-Biedl syndrome type 1, which has been mapped to 11q13. Received: 22 July 1998 / Accepted: 24 August 1998