Linkage and comparative mapping of the locus controlling susceptibility towards E. COLI F4ab/ac diarrhoea in pigs

In 1995, Edfors-Lilja and coworkers mapped the locus for the E. COLI K88ab (F4ab) and K88ac (F4ac) intestinal receptor to pig chromosome 13 (SSC13). Using the same family material we have refined the map position to a region between the microsatellite markers Sw207 and Sw225. Primers from these markers were used to screen a pig BAC library and the positive clones were used for fluorescent in situ hybridization (FISH) analysis. The results of the FISH analysis helped to propose a candidate gene region in the SSC13q41-->q44 interval. Shotgun sequencing of the FISH-mapped BAC clones revealed that the candidate region contains an evolutionary breakpoint between human and pig. In order to further characterise the rearrangements between SSC13 and human chromosome 3 (HSA3), detailed gene mapping of SSC13 was carried out. Based on this mapping data we have constructed a detailed comparative map between SSC13 and HSA3. Two candidate regions on human chromosome 3 have been identified that are likely to harbour the human homologue of the gene responsible for susceptibility towards E. COLI F4ab/ac diarrhoea in pigs.     

The GABAA receptor beta 3 subunit gene: characterization of a human cDNA from chromosome 15q11q13 and mapping to a region of conserved synteny on mouse chromosome 7.

A cDNA encoding the human GABAA receptor beta 3 subunit has been isolated from a brain cDNA library and its nucleotide sequence has been determined. This gene, GABRB3, has recently been mapped to human chromosome 15q11q13, the region deleted in Angelman and Prader-Willi syndromes. The association of distinct phenotypes with maternal versus paternal deletions of this region suggests that one or more genes in this region show parental-origin-dependent expression (genetic imprinting). Comparison of the inferred human beta 3 subunit amino acid sequence with beta 3 subunit sequences from rat, cow, and chicken shows a very high degree of evolutionary conservation. We have used this cDNA to map the mouse beta 3 subunit gene, Gabrb-3, in recombinant inbred strains. The gene is located on mouse chromosome 7, very closely linked to Xmv-33 between Tam-1 and Mtv-1, where two other genes from human 15q11q13 have also been mapped. This provides further evidence for a region of conserved synteny between human chromosome 15q11q13 and mouse chromosome 7. Proximal and distal regions of mouse chromosome 7 show genetic imprinting effects; however, the region of homology with human chromosome 15q11q13 has not yet been associated with these effects.     

Physical assignments of human chromosome 13 genes on pig chromosome 11 demonstrate extensive synteny and gene order conservation between pig and human

Previous mapping between the human and pig genomes suggested extensive conservation of human chromosome 13 (HSA13) to pig chromosome 11 (SSC11). The objectives of this study were comparative gene mapping of pig homologs of HSA13 genes and examining gene order within this conserved synteny group by physical assignment of each locus. A detailed HSA13 to SSC11 comparison was chosen since the comparative gene map is not well developed for these chromosomes and a rearranged gene order within conserved synteny groups was observed from the comparison between HSA13 and bovine chromosome 12 (BTA12). Heterologous primers for PCR were designed and used to amplify pig homologous fragments. The pig fragments were sequenced to confirm the homology. Six pig STSs (FLT1, ESD, RB1, HTR2A, EDNRB, and F10) were physically mapped using a somatic cell hybrid panel to SSC11, and fluorescent in situ hybridization (FISH) mapping was also applied to improve map resolution and determine gene order. Results from this study increase the comparative information available on SSC11 and suggest a conserved gene order on SSC11 and HSA13, in contrast to human:bovine comparisons of this syntenic group.     

The Human Obesity Gene Map: The 1998 Update

PÉRUSSE, LOUIS, YVON C. CHAGNON, JOHN WEISNAGEL, AND CLAUDE BOUCHARD. The human obesity gene map: the 1998 update. Obes Res. 1999;7:111–129. An update of the human obesity gene map incorporating published results up to the end of October 1998 is presented. Evidence from the human obesity cases caused by single gene mutations; other Mendelian disorders exhibiting obesity as a clinical feature; quantitative trait loci uncovered in human genome-wide scans and in crossbreeding experiments with mouse, rat, and pig models; association and case-control studies with candidate genes; and linkage studies with genes and other markers is reviewed. The most noticeable changes from the 1997 update is the number of obesity cases due to single gene mutations that increased from three cases due to mutations in two genes to 25 cases due to 12 mutations in seven genes. A look at the obesity gene map depicted in Figure 1 reveals that putative loci affecting obesity-related phenotypes are found on all but chromosome Y of the human chromosomes. Some chromosomes show at least three putative loci related to obesity on both arms (1, 2, 3, 6, 7, 8, 9, 11, 17, 19, 20, and X) and several on one chromosome arm only (4q, 5q, 10q, 12q, 13q, 15q, 16p, and 22q). The number of genes and other markers that have been associated or linked with human obesity phenotypes is increasing very rapidly and now approaches 27.     

The Human Obesity Gene Map: The 1997 Update

An update of the human obesity gene map incorporating published results up to October 1997 is presented. Evidence from Mendelian disorders exhibiting obesity as a clinical feature; single-gene mutation rodent models; quantitative trait loci uncovered in human genome-wide scans and in crossbreeding experiments with mouse, rat, and pig models; association and case-control studies with candidate genes; and linkage studies with genes and other markers is reviewed. All chromosomal locations of the animal loci are converted into human genome locations based on syntenic relationships between the genomes. A complete listing of all of these loci reveals that all but chromosome Y of the 24 human chromosomes are represented. Some chromosomes show at least three putative loci related to obesity on both arms (1, 2, 6, 8, 11, and 20) and several on one chromosome arm only (3p, 4q, 5q, 7q, 12q, 13q, 15q, 15p, 22q, and Xq). Studies reporting negative association and linkage results are also listed, with the exception of the unlinked markers from genome-wide scans.     

The telomeric region of BTA18 containing a potential QTL region for health in cattle exhibits high similarity to the HSA19q region in humans

We have applied a targeted physical mapping approach, based on the isolation of bovine region-specific large-insert clones using homologous human sequences and chromosome microdissection, to enhance the physical gene map of the telomeric region of BTA18 and to prove its evolutionary conservation. The latter is a prerequisite to exploit the dense human gene map for future positional cloning approaches. Partial sequencing and homology search were used to characterize 20 BACs targeted to the BTA18q2.4-q2.6 region. We used fluorescence in situ hybridization (FISH) to create physical maps of 11 BACs containing 15 gene loci; these BACs served as anchor loci. Using these approaches, 12 new gene loci (CKM, STK13, PSCD2, IRF3, VASP, ACTN4, ITPKC, CYP2B6, FOSB, DMPK, MIA, SIX5) were assigned on BTA18 in the bovine cytogenetic map. A resolved physical map of BTA18q2.4-q2.6 was developed, which encompasses 28 marker loci and a comparative cytogenetic map that contains 15 genes. The mapping results demonstrate the high evolutionary conservation between the telomeric region of BTA18q and HSA19q.     

Human homeo box-containing genes located at chromosome regions 2q31----2q37 and 12q12----12q13

Four human homeo box-containing cDNAs isolated from mRNA of an SV40-transformed human fibroblast cell line have been regionally localized on the human gene map. One cDNA clone, c10, was found to be nearly identical to the previously mapped Hox-2.1 gene at 17q21. A second cDNA clone, c1, which is 87% homologous to Hox-2.2 at the nucleotide level but is distinct from Hox-2.1 and Hox-2.2, also maps to this region of human chromosome 17 and is probably another member of the Hox-2 cluster of homeo box-containing genes. The third cDNA clone, c8, in which the homeo box is approximately 84% homologous to the mouse Hox-1.1 homeo box region on mouse chromosome 6, maps to chromosome region 12q12----12q13, a region that is involved in chromosome abnormalities in human seminomas and teratomas. The fourth cDNA clone, c13, whose homeo box is approximately 73% homologous to the Hox-2.2 homeo box sequence, is located at chromosome region 2q31----q37. The human homeo box-containing cluster of genes at chromosome region 17q21 is the human cognate of the mouse homeo box-containing gene cluster on mouse chromosome 11. Other mouse homeo box-containing genes of the Antennapedia class (class I) map to mouse chromosomes 6 (Hox-1, proximal to the IgK locus) and 15 (Hox-3). A mouse gene, En-1, with an engrailed-like homeo box (class II) and flanking region maps to mouse chromosome 1 (near the dominant hemimelia gene). Neither of the class I homeo box-containing genes--c8 and c13--maps to a region of obvious homology to chromosomal positions of the presently known mouse homeo box-containing genes.     

Improving the comparative map of SSC2p-q13 by sample sequencing of BAC clones

To improve the comparative map for pig chromosome 2 and increase the gene density on this chromosome, a porcine bacterial artificial chromosome (BAC) library was screened with 17 microsatellite markers and 18 genes previously assigned to pig chromosome 2. Fifty-one BAC clones located in the region of a maternally imprinted quantitative trait locus for backfat thickness (BFT) were identified. From these BACs 372 kb were sample sequenced. The average read length of a subclone was 442 basepair (bp). Contig assembly analysis showed that every bp was sequenced 1.28 times. Subsequently, sequences were compared with sequences in the nucleotide databases to identify homology with other mammalian sequences. Sequence identity was observed with sequences derived from 35 BACs. The average percentage identity with human sequences was 87.6%, with an average length of 143 bp. In total, sample sequencing of all BACs resulted in sequence identity with 29 human genes, 13 human expressed sequence tags (ESTs), 17 human genomic clones, one rat gene, one porcine gene and nine porcine ESTs. Eighteen genes located on human chromosome 11 and 19, and seven genes from other human locations, one rat gene and one porcine gene were assigned to pig chromosome 2 for the first time. The new genes were added to the radiation hybrid map at the same position as the locus from which the BAC that was sequenced was derived. In total 57 genes were placed on the radiation hybrid map of SSC2p-q13.     

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.     

Zoo-FISH with microdissected arm specific paints for HSA2, 5, 6, 16, and 19 refines known homology with pig and horse chromosomes

Microdissected arm specific paints (ASPs) for human (HSA) chromosomes (Chrs) 2, 5, 6, 16, and 19 were used as probes on pig (SSC) and horse (ECA) metaphase chromosomes. Regions homologous to individual human arms were delineated in the two species studied. Of the ten ASPs used, HSA6 and 16 ASPs showed complete synteny conservation of individual arms as single blocks/arms both in pig and horse. A similar trend was, in general, also observed for HSA19 ASPs. However, contrary to these observations, synteny conservation of individual arms of HSA2 and HSA5 was not observed in pig and horse. The arm specific painting data, coupled with the available gene mapping data, showed that, although HSA2 corresponded to two arms/chromosomes each in pig and horse, the breakpoint of this synteny in humans was not located at the centromere, but at HSA2q13 band. Similarly, arm specific paints for HSA5 showed that of the two blocks/chromosomes painted in pig and horse, one corresponded to HSA5q13-pter, the other to HSA5q13-qter. The findings suggest that 5q13 band may also be an evolutionary break point, similar to the one detected on HSA2q13. The microdissected human arm specific painting probes used in the present work provide more accurate and refined comparative information on pig and horse chromosomes than that available through the use of human whole chromosome specific paints. Received: 1 June 1997 / Accepted: 5 September 1997     

Radiation hybrid comparative mapping between human chromosome 17 and porcine chromosome 12 demonstrates conservation of gene order

A comparative study of human chromosome 17 (HSA17) and pig chromosome 12 (SSC12) was conducted using both somatic cell hybrid panel (SCHP) and radiation hybrid (RH) panel analysis. Sequences from an expressed sequence tag (EST) project in pig reproduction were examined and six genes and ESTs originally believed to map to HSA17 were selected for this study. The genes/ESTs were TATA box binding protein-associated factor (TAF2N/RBP56), alpha-2-plasmin inhibitor (SERPINF2/PLI), H3 histone family 3B (H3F3B), aminopeptidase puromycin sensitive (NPEPPS), an expressed sequence tag (ESTMI015) and P311 protein (P311). The SCHP analysis mapped five genes/ESTs (TAF2N, H3F3B, SERPINF2, NPEPPS and ESTMI015) to SSC12q11-q15 and SSC12p11-p15 with 100% concordance, and assigned P311 to SSC2 (1/2q24)-q29 with 100% concordance. Radiation hybrid analysis of all six genes confirmed the SCHP mapping results, with average retention frequency of 25%. Recent human sequence data demonstrated that P311 is actually located on HSA5q. As HSA5q and SSC2q show conserved syntenic regions predicted from bi-directional painting, our P311 mapping data is consistent with these results. An expanded comparative SSC12 RH map integrating the five new type I markers and 23 previously mapped loci was established using a LOD score threshold of 4.8. The gene order of the five genes/ESTs on the SSC12 framework RH map (H3F3B-ESTMI015-NPEPPS-TAF2N-SERPINF2) is identical to the HSA17 GB4 map but with inversion of the map as conventionally drawn.     

Mapping of AKT3, encoding a member of the Akt/protein kinase B family, to human and rodent chromosomes by fluorescence in situ hybridization

Previously, a rodent cDNA encoding the third member of the Akt/PKB family of serine/threonine kinases was cloned. We have now cloned the human homolog of this cDNA, and we have used this clone to map the AKT3 gene to human chromosome 1q44 by fluorescence in situ hybridization (FISH). We have also mapped the rodent homologs of AKT3 to rat chromosome 13q24-->q26 and mouse chromosome 1H4-6 by FISH.     

用放射杂交板定位鸡的MC4R基因以及其在鸡和人染色体上同源区的比较分析

黑素皮质素受体(melanocortin-4 receptor,MC4R)基因的突变与猪、鼠和人等的食欲、肥胖和生长有关联性,然而对鸡的MC4R基因的功能却知之甚少。为了确定鸡的MC4R基因在染色体上的位置,使用鸡-仓鼠杂交板(ChickRH6)做了该基因的定位工作。通过扩增ChickRH6杂交板上的93个样品,然后经整合分析将mC4R基因定位在2号染色体上的标记MCW0062、BCL2和OVY附近,即2q12。这个连锁图上的5个标记基于两点分析与MC4R的LOD值都大于5。同时,以MC4R基因为标记做了鸡和人的染色体比较分析。结果显示鸡的2号染色体(GGA2)和人的18号染色体(HSA18)存在同源区,且基因BCL2和肥胖基因(obesity)位于MC4R基因附近。推测鸡的MC4R基因与人的MC4R基因可能具有相似的功能。该研究揭示了鸡和人MC4R基因的染色体分布,并用杂交放射板将鸡的MC4R基因定位在2号染色体的12区带。     

Generation of a 5.5-Mb BAC/PAC contig of pig chromosome 6q1.2 and its integration with existing RH, genetic and comparative maps

We generated a sequence-ready BAC/PAC contig spanning approximately 5.5 Mb on porcine chromosome 6q1.2, which represents a very gene-rich genome region. STS content mapping was used as the main strategy for the assembly of the contig and a total of 6 microsatellite markers, 53 gene-related STS and 116 STS corresponding to BAC and PAC end sequences were analyzed. The contig comprises 316 BAC and PAC clones covering the region between the genes GPI and LIPE. The correct contig assembly was verified by RH-mapping of STS markers and comparative mapping of BAC/PAC end sequences using BLAST searches. The use of microsatellite primer pairs allowed the integration of the physical maps with the genetic map of this region. Comparative mapping of the porcine BAC/PAC contig with respect to the gene-rich region on the human chromosome 19q13.1 map revealed a completely conserved gene order of this segment, however, physical distances differ somewhat between HSA19q13.1 and SSC6q1.2. Three major differences in DNA content between human and pig are found in two large intergenic regions and in one region of a clustered gene family, respectively. While there is a complete conservation of gene order between pig and human, the comparative analysis with respect to the rodent species mouse and rat shows one breakpoint where a genome segment is inverted.     

Chromosomal localization of the human and mouse histidine decarboxylase genes by in situ hybridization. Exclusion of the HDC gene from the Prader-Willi syndrome region

Using a rat histidine decarboxylase (HDC) cDNA probe, we have mapped the HDC gene by in situ hybridization to the ql5–q2l region of human chromosom e15 and to the E5-G region of murine chromosome 2. These localizations strengthen a syntenic group conserved between human chromosome 15 and mouse chromosome 2. The localization of the HDC gene on the human chromosome 15 map shows that it is not included within the Prader-Willi Syndrome region (PWCR).     

Juxta-centromeric region of human chromosome 21 is enriched for pseudogenes and gene fragments

A physical map including four pseudogenes and 10 gene fragments and spanning 500 kb in the juxta-centromeric region of the long arm of human chromosome 21 is presented. cDNA fragments isolated from a selected cDNA library were characterized and mapped to the 831B6 YAC and to two BAC contigs that cover 250 kb of the region. An 85 kb genomic sequence located in the proximal region of the map was analyzed for putative exons. Four pseudogenes were found, including psiIGSF3, psiEIF3, psiGCT-rel whose functional copies map to chromosome 1p13, chromosome 2 and chromosome 22q11, respectively. The TTLL1 pseudogene corresponds to a new gene whose functional copy maps to chromosome 22q13. Ten gene fragments represent novel sequences that have related sequences on different human chromosomes and show 97-100% nucleotide identity to chromosome 21. These may correspond to pseudogenes on chromosome 21 and to functional genes in other chromosomes. The 85 kb genomic sequence was analyzed also for GC content, CpG islands, and repetitive sequence distribution. A GC-poor L isochore spanning 40 kb from satellite 1 was observed in the most centromeric region, next to a GC-rich H isochore that is a candidate region for the presence of functional genes. The pericentric duplication of a 7.8 kb region that is derived from the 22q13 chromosome band is described. We showed that the juxta-centromeric region of human chromosome 21 is enriched for retrotransposed pseudogenes and gene fragments transferred by interchromosome duplications, but we do not rule out the possibility that the region harbors functional genes also.     

Autistic disorder and chromosome 15q11-q13: construction and analysis of a BAC/PAC contig

Autistic disorder (AD) is a neurodevelopmental disorder that affects approximately 2-10/10,000 individuals. Chromosome 15q11-q13 has been implicated in the genetic etiology of AD based on (1) cytogenetic abnormalities; (2) increased recombination frequency in this region in AD versus non-AD families; (3) suggested linkage with markers D15S156, D15S219, and D15S217; and (4) evidence for significant association with polymorphisms in the gamma-aminobutyric acid receptor subunit B3 gene (GABRB3). To isolate the putative 15q11-q13 candidate AD gene, a genomic contig and physical map of the approximately 1.2-Mb region from the GABA receptor gene cluster to the OCA2 locus was generated. Twenty-one bacterial artificial chromosome (BAC) clones, 32 P1-derived artificial chromosome (PAC) clones, and 2 P1 clones have been isolated using the markers D15S540, GABRB3, GABRA5, GABRG3, D15S822, and D15S217, as well as 34 novel markers developed from the end sequences of BAC/PAC clones. In contrast to previous findings, the markers D15S822 and D15S975 have been localized within the GABRG3 gene, which we have shown to be approximately 250 kb in size. NotI and numerous EagI restriction enzyme cut sites were identified in this region. The BAC/PAC genomic contig can be utilized for the study of genomic structure and the identification and characterization of genes and their methylation status in this autism candidate gene region on human chromosome 15q11-q13.     

Mapping four genes from human chromosome 4 to porcine chromosome 8 further develops the comparative map for an economically important chromosome of the swine genome

Because porcine chromosome (SSC) 8 has become the focal point of many efforts aimed at identifying quantitative trait loci affecting ovulation rate, genes distributed across human chromosome (HSA) 4 were physically mapped in the pig. A more refined comparative map of this region for these two species was produced. In this study, four genes were selected based on their location in the human genome, the availability of nucleotide sequence and their genomic organization. The genes selected were fibroblast growth factor basic (FGF2; HSA 4q25-27), gonadotropin releasing hormone receptor (GNRHR; HSA 4q13), phosphodiesterase 6 B (PDE6B; HSA 4p16.3) and aminopeptidase S (PEPS; HSA 4p11-q12). Genomic libraries were screened via PCR and clones were physically assigned using fluorescence in situ hybridization (FISH). These four genes from HSA 4 were physically mapped to SSC 8p2.3 (PDE6B), 8p1.1 (PEPS), 8q1.1-1.2 (GNRHR) and 8q2.2-2.4 (FGF2). These assignments provide additional benchmarks for the comparative map and help define the level of gene order conserved between HSA 4 and SSC 8.     

Chromosomal location of murine and human IL-1 receptor genes.

The gene for the type I interleukin-1 (IL-1) receptor has been mapped in both mouse and human. In the human genome, a combination of segregation analysis of rodent-human hybrid cells and chromosomal in situ hybridization has placed the gene on the long arm of chromosome 2, at band 2q12. This is near the reported map position of the loci for IL-1 alpha and IL-1 beta (2q13----2q21). The murine gene has been mapped by analysis of restriction fragment length polymorphisms in interspecific backcrosses to the centromeric end of chromosome 1, in a region that is syntenic to a portion of human chromosome 2. The murine Il-1r1 gene has thus been separated from the IL-1 genes, which lie on murine chromosome 2.