Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Cosmids”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,117 records · Page 62Linked to original sources
PubMed1994

Subchromosomal band interval mapping and ordering of DNA markers in the region 3q26.3-q27 involved in the dup(3q) syndrome.

The duplication 3q syndrome is characterized by the partial trisomy of a segment of the long arm of chromosome 3. This segment, although variable in size, includes 3q26.3-q27 as the minimal region of overlap. We have previously used patient chromosome breakpoints to select cosmids within this region. In this report, we have used two- and three-color fluorescence in situ hybridization on metaphase and interphase chromosomes to perform high-resolution cytological mapping of the six cosmids identified. The results allowed us to determine the centromere-telomere orientation, the order, and the relative distances of the markers used. Because some of the markers used are part of the consensus chromosome 3 map, our data can be easily integrated with existing mapping information about this chromosome. Our data provide a framework for further physical mapping studies of this region.

Abnormalities, Multiple↗
PubMed1995

Isolation and mapping of 186 new DNA markers on human chromosome 1.

To obtain DNA markers on human chromosome 1, we first isolated 500 cosmid clones from mouse A9 cells containing a human chromosome 1 tagged with pSV2neo. Of these, 186 were localized on each band of human chromosome 1 by R-banding fluorescence in situ hybridization; 118 and 68 were on the short and long arms, respectively. We performed restriction fragment length polymorphism (RFLP) analysis of these cosmid clones, and polymorphism was recognized with one or more enzyme in 43 of them. Two markers proved to have variable numbers of tandem repeats. Since several tumor suppressor genes, as well as genes responsible for hereditary disorders, may be located on this human chromosome, the DNA markers will be useful for RFLP analysis or the isolation of new genes related to various disorders.

Chromosome Mapping↗
PubMed1995

A high-resolution map of genes, microsatellite markers, and new dinucleotide repeats from UBE1 to the GATA locus in the region Xp11.23.

Several new genes and markers have recently been identified on the proximal short arm of the human X chromosome in the area of Xp11.23. We had previously generated a YAC contig in this region extending from UBE1 to the OATL1 locus. In this report two polymorphic dinucleotide repeats, DXS6949 and DXS6950, were isolated and characterized from the OATL1 locus. A panel of YAC deletion derivatives from the distal portion of the contig was used in conjunction with the rest of the YAC map to position the new microsatellites and order other markers localizing to this interval. The marker order was determined to be DXS1367-ZNF81-DXS6849-ZNF21-DXS6616-DXS 6950-DXS6949. In the proximal region below OATL1, we have isolated a pair of YACs from the GATA locus, B1026 and C01160. Mapping within these YACs indicates the orientation of DXS1126 and DXS1240, while a cosmid near the OATL1 region reveals the overlap between the YAC contigs from the two loci. This cosmid contains the gene responsible for Wiskott-Aldrich syndrome (WAS) and localizes the disease gene between OATL1 and GATA. These data enable the expansion of the present physical map of the X chromosome from UBE1 to the GATA locus, covering a large portion of the Xp11.23 region. Genetic cross-overs in Xp11.23 support the marker orientation and the position of WAS, contrary to previous reports. With the integration of both physical and genetic maps we have predicted the following marker order: Xpter-UBE1-SYN1/ARAF1/ TIMP1-DXS1367-ZNF81-DXS.6849-ZNF21-DXSy6616++ +-(OATL1, DXS6950-DXS6949)- WAS-(GATA, DXS1126)-DXS1240-Xcen.(ABSTRACT TRUNCATED AT 250 WORDS)

Chromosome Mapping↗
PubMed1995

The human replacement histone H3.3B gene (H3F3B).

H3.3 is a replacement histone subtype that is encoded by two replication-independent genes termed H3.3A and H3.3B, respectively. We have isolated a fullsize H3.3 cDNA clone from an oligo(dT)-primed human testicular cDNA library. Subsequently, the corresponding gene was isolated from a human cosmid library and was identified as the H3.3B gene. It was the only histone gene on this 42-kb cosmid clone. The gene structure shows characteristic features of an H3.3 gene. First, it contains an intron of about 0.5 kb in the 5' untranslated region and two smaller introns within the coding gene portion. Second, no histone gene-specific dyad symmetry element was found in the 3' untranslated region, but three putative polyadenylation signals were detected downstream of the gene. The corresponding transcripts were detected by Northern blot analysis using poly(A)+ RNA from testis and from the HEK293 tumor cell line. The newly discovered human H3.3B gene (HGMW-approved symbol H3F3B) was mapped by fluorescence in situ hybridization to the telomeric region of chromosome 17 (17q25). This localization of the H3.3B gene and its solitary arrangement contrast with the majority of the replication-dependent histone genes, which form a large cluster on chromosome 6 and a second cluster on chromosome 1.

Amino Acid Sequence↗
PubMed1995

Isolation of murine telomere-proximal sequences by affinity capture and PCR.

We describe a method of selectively enriching for murine telomere-proximal sequences using affinity capture followed by PCR amplification. The telomeric fragments were selected from NotI-digested and lambda exonuclease-resected mouse genomic DNA by annealing to a biotinylated riboprobe containing multiple copies of the telomere repeat (TTAGGG)n. The resultant DNA-RNA hybrids were selectively retained on a matrix with covalently bound avidin. The captured DNA was then specifically released by ribonuclease action, and PCR amplification was performed using mouse repeat primers. The PCR products were cloned and used to screen a mouse genomic cosmid library, and the resultant cosmid clones were analyzed by fluorescence in situ hybridization. Ten of 70 clones analyzed gave telomere-proximal hybridization signals, indicating an at least 500-fold enrichment for telomere-proximal sequences.

Animals↗
PubMed1995

Cloning of a highly conserved human protein serine-threonine phosphatase gene from the glioma candidate region on chromosome 19q13.3.

Allelic loss studies have suggested that a glioma tumor suppressor gene resides in a 425-kb region of chromosome 19q, telomeric to D19S219 and centromeric to D19S112. Exon amplification of a cosmid contig spanning this region yielded four exons with high homology to a rat protein serine-threonine phosphatase from a cosmid approximately 100 kb telomeric to D19S219. Isolation of a near full-length cDNA from a human fetal brain cDNA library revealed a protein serine-threonine phosphatase with a tetratricopeptide motif, almost identical to human PPP5C (PP5) and highly homologous to rat PPT. Northern blotting demonstrated expression in most tissues, including brain. Primary and cultured gliomas were studied for genetic alterations in this gene using pulsed-field gel electrophoresis, routine Southern blots, and genomic DNA-and RNA-based single-strand conformation polymorphism analysis. Genomic alterations were were not detected in any of the gliomas, and all studied gliomas expressed the gene, suggesting that this phosphatase is not the putative chromosome 19q glioma tumor suppressor gene.

Amino Acid Sequence↗
PubMed1996

High-resolution mapping of a 130-kb core region of the MYCN amplicon in neuroblastomas.

The MYCN proto-oncogene is amplified in 25% of neuroblastomas, and amplification is strongly correlated with advanced disease stage and with rapid tumor progression. We have generated a high-resolution restriction map of nearly 500 kb spanning the MYCN locus by subcloning yeast artificial chromosomes into cosmids. Cosmids plus additional amplified probes were hybridized to DNA from 33 MYCN amplified neuroblastomas, and we determined that the amplicons range from 350 kb to over 1 Mb. Deletions and rearrangements of the MYCN amplicon occurred less frequently in primary tumors than in cell lines. We have defined a 130-kb region that was amplified in 32 of 33 tumors. One additional tumor deleted 65 kb of this region from its amplicon, while amplifying a large amount of flanking DNA. The only CpG island found in this region was within the MYCN gene. In conclusion, our data demonstrate that, despite the large size of most amplicons, the core domain that is consistently amplified in neuroblastomas probably contains little more than the MYCN gene.

Chromosomes, Artificial, Yeast↗
PubMed1996

Structural analysis of the HLA-A/HLA-F subregion: precise localization of two new multigene families closely associated with the HLA class I sequences.

Positional cloning strategies for the hemochromatosis gene have previously concentrated on a target area restricted to a maximum genomic expanse of 400 kb around the HLA-A and HLA-F loci. Recently, the candidate region has been extended to 2-3 Mb on the distal side of the MHC. In this study, 10 coding sequences [hemochromatosis candidate genes (HCG) I to X] were isolated by cDNA selection using YACs covering the HLA-A/HLA-F subregion. Two of these (HCG II and HCG IV) belong to multigene families, as well as other sequences already described in this region, i.e., P5, pMC 6.7, and HLA class 1. Fingerprinting of the four YACs overlapping the region was performed and allowed partial localization of the different multigene family sequences on each YAC without defining their exact positions. Fingerprinting on cosmids isolated from the ICRF chromosome 6-specific cosmid library allowed more precise localization of the redundant sequences in all of the multigene families and revealed their apparent organization in clusters. Further examination of these intertwined sequences demonstrated that this structural organization resulted from a succession of complex phenomena, including duplications and contractions. This study presents a precise description of the structural organization of the HLA-A/HLA-F region and a determination of the sequences involved in the megabase size polymorphism observed among the A3, A24, and A31 haplotypes.

Cell Line↗
PubMed1996

A 2-megabase physical contig incorporating 43 DNA markers on the human X chromosome at p11.23-p11.22 from ZNF21 to DXS255.

A comprehensive physical contig of yeast artificial chromosomes (YACs) and cosmid clones between ZNF21 and DXS255 has been constructed, spanning 2 Mb within the region Xp11.23-p11.22. As a portion of the region was found to be particularly unstable in yeast, the integrity of the contig is dependent on additional information provided by the sequence-tagged site (STS) content of cosmid clones and DNA marker retention in conventional and radiation hybrids. The contig was formatted with 43 DNA markers, including 19 new STSs from YAC insert ends and an internal Alu-PCR product. The density of STSs across the contig ranges from one marker every 20 kb to one every 60 kb, with an average density of one marker every 50 kb. The relative order of previously known genes and expressed sequence tags in this region is predicted to be Xpter-ZNF21-DXS7465E (MG66)-DXS7927E (MG81)-WASP, DXS1011E, DXS7467E (MG21)-DXS- 7466E (MG44)-GATA1-DXS7469E (Xp664)-TFE3-SYP (DXS1007E)-Xcen. This contig extends the coverage in Xp11 and provides a framework for the future identification and mapping of new genes, as well as the resources for developing DNA sequencing templates.

Animals↗
PubMed1996

The human B22 subunit of the NADH-ubiquinone oxidoreductase maps to the region of chromosome 8 involved in branchio-oto-renal syndrome.

To identify candidate genes for Branchio-oto-renal (BOR) syndrome, we have made use of a set of cosmids that map to 8q13.3, which has previously been shown to be involved in this syndrome. These cosmids were used as genomic clones in the attempts to isolate corresponding cDNAs using a modified hybrid selection technique. cDNAs from the region were identified and used to search for sequence similarity in human or other species. One cDNA clone was found to have 89% sequence similarity to the bovine B22 subunit of NADH-ubiquinone oxidoreductase, a mitochondrial protein in the respiratory electron transport chain. Given the history of other mitochondrial mutations being involved in hearing loss syndromes, this gene should be considered a strong candidate for involvement in BOR.

Abnormalities, Multiple↗
PubMed1996

Physical mapping of receptor type protein tyrosine phosphatase sigma (PTPRS) to human chromosome 19p13.3.

Receptor type protein tyrosine phosphatase sigma (PTPRS), a member of a subfamily of receptor type protein tyrosine phosphatases, has been mapped to human chromosome 19p13.3 using fluorescence in situ hybridization (FISH) analysis. Hybridization analysis of chromosome 19 library cosmids has revealed several positive clones that are part of a contig located in the same region. The location of this contig was verified since one of the cosmid clones is anchored to a high-resolution FISH map of human chromosome 19p. In addition, the location of this gene relative to previously mapped proximal markers reveals a new point in the human-mouse synteny map by extending the mouse chromosome 17 synteny region in the telomeric direction.

Animals↗
PubMed1996

A 1.5-megabase physical map encompassing the multiple endocrine neoplasia type-1 (MEN1) locus on chromosome 11q13.

Linkage analysis and loss of heterozygosity studies have shown that the gene responsible for the multiple endocrine neoplasia type-1 (MEN1) syndrome localizes to a small interval between D11S427 and D11S460 on chromosome 11q13. As an initial step to clone this tumor suppressor gene, our group is the first to map the MEN1 region physically using yeast artificial chromosome, bacterial artificial chromosome (BAC), and cosmid contigs. The 1.5-Mb high-resolution, contiguous map extends from PYGM to 300 kb telomeric of D11S460. Of this, the 1.2-Mb interval between PYGM and D11S460 is isolated in cosmids and BACs and will be useful for the development of genomic sequences and transcription maps of this important region. Nine new sequence-tagged sites (STS) are also characterized from this region. The physical map and the STSs will be valuable tools for the cloning of the MEN1 gene.

Chromosome Mapping↗
PubMed1996

Isolation and characterization of GT335, a novel human gene conserved in Escherichia coli and mapping to 21q22.3.

As part of efforts to identify candidate genes for disorders mapped to 21q22.3, we have constructed a 405-kb cosmid contig encompassing five tightly linked markers mapping to this region. A subset of these cosmids was used to identify cDNA fragments by the method of hybrid selection. We present here the cDNA sequence of one such gene (GT335) mapping to this region. The gene is expressed as a 1.7-kb transcript predominantly in heart and skeletal muscle, potentially displays alternate splicing, and is predicted to encode a protein 268 amino acids in length. GT335 spans an estimated 13 kb of genomic DNA and is split into seven exons. Five of the six introns conform to the GT . . . AG consensus for intronic splice junctions; the sixth contains nonconventional (AT . . . AC) intronic junctions. We screened this gene for single-basepair mutations using single-strand conformation polymorphism and sequence analysis of both cDNA and genomic DNA from a number of unrelated individuals and have identified several sequence variations, two of which cause conservative amino acid substitutions. This gene is well conserved evolutionarily, with homologs identified in zebrafish and Escherichia coli, suggesting that it plays an important role in basic cellular metabolism.

Amino Acid Sequence↗
PubMed1997

Physical mapping 220 kb centromeric of the human MHC and DNA sequence analysis of the 43-kb segment including the RING1, HKE6, and HKE4 genes.

A cosmid contig was constructed from a YAC clone with a 220-kb insert that spans the centromeric side of the human MHC class II region, corresponding to the mouse t complex. The gene order was identified to be HSET-HKE1.5-HKE2-HKE3-RING1-HKE6- HKE4 (RING5). The genomic sequence of a 42,801-bp long region encoded by one cosmid clone in the RING1, HKE6, and HKE4 subregions was determined by the shotgun method. The exon-intron organization of these three genes, RING1 (Ring finger protein), HKE6 (steroid dehydrogenase-like protein), and HKE4 (transmembrane protein with histidine-rich charge clusters), was determined. The previously reported RING2 gene was revealed to be identical to HKE6. Transcripts from HKE4 were detected in the placenta, lung, kidney, and pancreas. Those of HKE6 were found in the liver and pancreas. The 25-kb region proximal to the RING1 gene includes an extensive dense cluster of Alu repeats (about 1.2 Alu per kb), and no gene has been identified in this so far. The region is equivalent to part of the mouse t complex and could be of relevance to human development.

Amino Acid Sequence↗
PubMed1997

Construction of a 1.2-Mb sequence-ready contig of chromosome 11q13 encompassing the multiple endocrine neoplasia type 1 (MEN1) gene. The European Consortium on MEN1.

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant familial cancer syndrome characterized by parathyroid, pancreatic, and anterior pituitary tumors. The MEN1 locus has been previously localized to chromosome 11q13, and a 2-Mb gene-rich region flanked by D11S1883 and D11S449 has been defined. We have pursued studies to facilitate identification of the MEN1 gene by narrowing this critical region to a 900-kb interval between the VRF and D11S1783 loci through melotic mapping. This was achieved by investigating 17 cosmids for microsatellite polymorphisms, which defined two novel polymorphisms at the VRF and A0138 loci, and utilizing these to characterize recombinants in MEN1 families. In addition, we have established a 1200-kb sequence-ready contig consisting of 26 cosmids, eight BACs, and eight PACs that encompass this region. The precise locations for 19 genes and three ESTs within this contig have been determined, and three gene clusters consisting of a centromeric group (VRF, FKBP2, PNG, and PLCB3), a middle group (PYGM, ZFM1, SCG1, SCG2 (which proved to be the MEN1 gene), and PPP2R5B), and a telomeric group (H4B, ANG3, ANG2, ANG1, FON, FAU, NOF, NON, and D11S2196E) were observed. These results represent a valuable transcriptional map of chromosome 11q13 that will help in the search for disease genes in this region.

Chromosome Mapping↗
PubMed1997

Sequence-based exon prediction around the synaptophysin locus reveals a gene-rich area containing novel genes in human proximal Xp.

The human Xp11.23-p11.22 interval has been implicated in several inherited diseases including Wiskott-Aldrich syndrome; three forms of X-linked hypercalciuric nephrolithiaisis; and the eye disorders retinitis pigmentosa 2, congenital stationary night blindness, and Aland Island eye disease. In constructing YAC contigs spanning Xp11. 23-p11.22, we have previously shown that the region around the synaptophysin (SYP) gene is refractory to cloning in YACs, but highly stable in cosmids. Preliminary analysis of the latter suggested that this might reflect a high density of coding sequences and we therefore undertook the complete sequencing of a SYP-containing cosmid. Sequence data were extensively analyzed using computer programs such as CENSOR (to mask repeats), BLAST (for homology searches), and GRAIL and GENE-ID (to predict exons). This revealed the presence of 29 putative exons, organized into three genes, in addition to the 7 exons of the complete SYP coding region, all mapping within a 44-kb interval. Two genes are novel, one (CACNA1F) showing high homology to alpha1 subunits of calcium channels, the other (LMO6) encoding a product with significant similarity to LIM-domain proteins. RT-PCR and Northern blot studies confirmed that these loci are indeed transcribed. The third locus is the previously described, but not previously localized, A4 differentiation-dependent gene. Given that the intron-exon boundaries predicted by the analysis are consistent with previous information where available, we have been able to suggest the genomic organization of the novel genes with some confidence. The region has an elevated GC content (>53%), and we identified CpG islands associated with the 5' ends of SYP, A4, and LMO6. The order of loci was Xpter-A4-LMO6-SYP-CACNA1F-Xcen, with intergenic distances ranging from approximately 300 bp to approximately 5 kb. The density of transcribed sequences in this area (>80%) is comparable to that found in the highly gene-rich chromosomal band Xq28. Further studies may aid our understanding of the long-range organization surrounding such gene-enriched regions.

Amino Acid Sequence↗
PubMed1998

Transcriptional map of the 2.5-Mb CBR-ERG region of chromosome 21 involved in Down syndrome.

The region of chromosome 21 between genes CBR and ERG (CBR-ERG region), which spans 2.5 Mb on 21q22.2, has been defined by analysis of patients with partial trisomy 21. It contributes significantly to the pathogenesis of many characteristics of Down syndrome, including morphological features, hypotonia, and mental retardation. Cosmid contigs covering 80% of the region were constructed and EcoRI maps produced. These cosmids were used for exon trapping and cDNA selection from three cDNA libraries (fetal brain, fetal liver, and adult skeletal muscle). Isolated exons and cDNAs were mapped on the EcoRI map, organized into contigs, sequenced, and used as probes for Northern blot analysis of RNA from fetal and adult tissues. We identified 27 genuine or highly probable transcriptional units evenly distributed along the CBR-ERG region. Eight of the transcriptional units are known genes.

Chromosome Mapping↗
PubMed1995

GABA receptor minigene rescues insecticide resistance phenotypes in Drosophila.

A single point mutation within the GABA receptor gene Resistance to dieldrin (Rdl) confers a high level of resistance to cyclodiene insecticides in a wide range of insects. Previous studies have shown partial rescue of the susceptible phenotype via germline transformation of a 36 kb cosmid coding (or all four alternative Rdl splice forms. Here, we describe the construction of two Rdl promoter/cDNA minigenes, each coding for one of the splice forms alone. Single splice forms rescued both the insecticide susceptible and resistant phenotypes associated with the locus as effectively as the complete cosmid. The minigenes also rescue the lethality associated with homozygous re-arrangements disrupting the Rdl gene, and the level of rescue observed is not increased by the addition of more than one splice form. This demonstrates that only a single Rdl splice form is necessary both to confer insecticide sensitivity and also to rescue lethality. Methods by which phenotype rescue could be enhanced and the potential advantages of using Rdl as a selectable marker are discussed.

Animals↗