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At least 1,621 records · Page 90Linked to original sources
PubMed1995

High-resolution physical mapping of a 250-kb region of human chromosome 11q24 by genomic sequence sampling (GSS).

A physical map of the region of human chromosome 11q24 containing the FLI1 gene, disrupted by the t(11;22) translocation in Ewing sarcoma and primitive neuroectodermal tumors, was analyzed by genomic sequence sampling. Using a 4- to 5-fold coverage chromosome 11-specific library, 22 region-specific cosmid clones were identified by phenol emulsion reassociation hybridization, with a 245-kb yeast artificial chromosome clone containing the FLI1 gene, and by directed "walking" techniques. Cosmid contigs were constructed by individual clone fingerprinting using restriction enzyme digestion and assembly with the Genome Reconstruction and AsseMbly (GRAM) computer algorithm. The relative orientation and spacing of cosmid contigs with respect to the chromosome was determined by the structural analysis of cosmid clones and by direct visual in situ hybridization mapping. Each cosmid clone in the contig was subjected to "one-pass" end sequencing, and the resulting ordered sequence fragments represent approximately 5% of the complete DNA sequence, making the entire region accessible by PCR amplification. The sequence samples were analyzed for putative exons, repetitive DNAs, and simple sequence repeats using a variety of computer algorithms. Based upon the computer predictions, Southern and Northern blot experiments led to the independent identification and localization of the FLI1 gene as well as a previously unknown gene located in this region of chromosome 11q24. This approach to high-resolution physical analysis of human chromosomes allows the assembly of detailed sequence-based maps and provides a tool for further structural and functional analysis of the genome.

Base Sequence↗
PubMed1997

Analysis of 1;17 translocation breakpoints in neuroblastoma: implications for mapping of neuroblastoma genes.

Deletions and translocations resulting in loss of distal 1p-material are known to occur frequently in advanced neuroblastomas. Fluorescence in situ hybridisation (FISH) showed that 17q was most frequently involved in chromosome 1p translocations. A review of the literature shows that 10 of 27 cell lines carry 1;17 translocations. Similar translocations were also observed in primary tumours. Together with the occurrence of a constitutional 1;17 translocation in a neuroblastoma patient, these observations suggest a particular role for these chromosome re-arrangements in the development of neuroblastoma. Apart from the loss of distal 1p-material, these translocations invariably lead to extra copies of 17q. This also suggested a possible role for genes on 17q in neuroblastoma tumorigenesis. Further support for this hypothesis comes from the observation that in those cell lines without 1;17 translocations, other chromosome 17q translocations were present. These too lead to extra chromosome 17q material. Molecular analysis of 1;17 translocation breakpoints revealed breakpoint heterogeneity both on 1p and 17q, which suggests the involvement of more than 2 single genes on 1p and 17q. The localisation of the different 1p-breakpoints occurring in 1;17 translocations in neuroblastoma are discussed with respect to the recently identified candidate tumor suppressor regions and genes on 1p. In this study, we focused on the molecular analysis of the 17q breakpoints in 1;17 translocations. Detailed physical mapping of the constitutional 17q breakpoint allowed for the construction of a YAC contig covering the breakpoint. Furthermore, a refined position was determined for a number of 17q breakpoints of 1;17 translocations found in neuroblastoma cell lines. The most distal 17q breakpoint was identified in cell line UHG-NP and mapped telomeric to cosmid cCI17-1049 (17q21). This suggests that genes involved in a dosage-dependent manner in the development of neuroblastoma map in the distal segment 17q22-qter. Future studies aim at the molecular cloning of 1;17 translocation breakpoints and at deciphering the mechanisms leading to 1;17 translocations and possibly to the identification of neuroblastoma genes at or in the vicinity of these breakpoints.

Chromosome Mapping↗
PubMed1996

Identification and mapping of type 1 neurofibromatosis (NF1) homologous loci.

During the establishment of a YAC contig for the type 1 neurofibromatosis (NF1) region on human chromosome 17q11.2, several YAC clones were isolated which originated from a different chromosome but which retained strong homology to NF1 coding regions (Marchuk et al., 1992). Fluorescence in situ hybridization (FISH) using these clones has identified NF1-homologous loci on several human chromosomes, including 2, 14, 15, 21, and 22. PCR amplification using primers originally designed to amplify NF1 exons has confirmed these chromosome localizations and, in addition, has revealed NF1-homologous sequences on chromosomes 12 and 20. Sequence analysis of the amplified products has demonstrated that (1) most of these loci have > 90% identity with NF1 sequences; (2) most of these loci represent nonprocessed pseudogenes; and (3) for the chromosome 12 locus, the two described regions of homology with NF1 have open reading frames. Finally, we have identified two novel alpha-satellite DNA repeat units in proximity to NF1-homologous sequences on chromosome 14. Their association suggests a mechanism for dispersion of the NF1-homologous loci based on alpha-satellite-mediated exchange between nonhomologous chromosomes.

Base Sequence↗
PubMed1996

Narrowing the critical region for a rhabdoid tumor locus in 22q11.

Rhabdoid tumor is a rare malignant neoplasm of childhood that may occur in various locations, including the central nervous system and the kidney. Previous cytogenetic studies of primary rhabdoid tumors have demonstrated monosomy or deletion of chromosome 22 and have implicated the presence of a rhabdoid tumor suppressor gene that maps to 22q. We have employed fluorescence in situ hybridization to narrow the region for this locus in four rhabdoid tumor cell lines with translocations or deletions involving chromosome segment 22q11. The completion of a cosmid and yeast artificial chromosome contig spanning the immunoglobulin lambda gene locus to BCR has allowed us to map a critical region for a rhabdoid tumor gene to a 500 kb span of chromosome segment 22q11.

Chromosome Mapping↗
PubMed2000

A sequence-ready map of the Usher syndrome type III critical region on chromosome 3q.

Usher syndrome type 3 (USH3; MIM 276902) is an autosomal recessive disorder associated with progressive hearing loss and retinal degeneration. We recently refined the localization of USH3 to a 1-cM genetic interval between markers D3S1299 and D3S3625. We have now constructed a bacterial artificial chromosome contig over the region. Novel polymorphic markers were generated and physically fine-mapped, allowing further narrowing of the critical interval to a 250-kb genomic fragment. Of seven ESTs mapping to the initial critical region, WI-11588 and SHGC-133 represent the human SIAH2 gene, which was excluded as a candidate for USH3 by sequencing and subsequently, by its position. KIAA0001 and D3S3882 derive from the transcript of a putative G-protein-coupled receptor gene that was excluded as a candidate by sequencing of patient DNA. These data provide a basis for the sequencing and final characterization of the USH3 region and isolation of the disease gene.

Chromosomes, Bacterial↗
PubMed1998

Two sequence-ready contigs spanning the two copies of a 200-kb duplication on human 21q: partial sequence and polymorphisms.

Physical mapping across a duplication can be a tour de force if the region is larger than the size of a bacterial clone. This was the case of the 170- to 275-kb duplication present on the long arm of chromosome 21 in normal human at 21q11.1 (proximal region) and at 21q22.1 (distal region), which we described previously. We have constructed sequence-ready contigs of the two copies of the duplication of which all the clones are genuine representatives of one copy or the other. This required the identification of four duplicon polymorphisms that are copy-specific and nonallelic variations in the sequence of the STSs. Thirteen STSs were mapped inside the duplicated region and 5 outside but close to the boundaries. Among these STSs 10 were end clones from YACs, PACs, or cosmids, and the average interval between two markers in the duplicated region was 16 kb. Eight PACs and cosmids showing minimal overlaps were selected in both copies of the duplication. Comparative sequence analysis along the duplication showed three single-basepair changes between the two copies over 659 bp sequenced (4 STSs), suggesting that the duplication is recent (less than 4 mya). Two CpG islands were located in the duplication, but no genes were identified after a 36-kb cosmid from the proximal copy of the duplication was sequenced. The homology of this chromosome 21 duplicated region with the pericentromeric regions of chromosomes 13, 2, and 18 suggests that the mechanism involved is probably similar to pericentromeric-directed mechanisms described in interchromosomal duplications.

Cell Line↗
PubMed2000

Molecular characterization of two soybean homologs of Arabidopsis thaliana CLAVATA1 from the wild type and fasciation mutant.

Arabidopsis thaliana CLAVATA1 (CLV1)-like genes were isolated from the wild type and fasciation mutant of soybean (Glycine max). Two soybean homologs of CLV1, designated GmCLV1A and GmCLV1B, are similar in sequence. No missense mutations in GmCLV1A and GmCLV1B between the wild type and mutant were found. GmCLV1 was mapped at 7. 1 cM from a restriction fragment length polymorphism marker on the linkage group H of the soybean molecular map. DNA fingerprinting using bacterial artificial chromosome clones identified two contigs indicating there are two loci for GmCLV1 in the soybean genome. One locus contains GmCLV1A and the other locus contains GmCLV1A and GmCLV1B. Relative multiplex quantitative reverse transcriptase-polymerase chain reaction analysis indicates that the two genes are transcribed in all organs and at higher levels in floral apices. Differential expression patterns of the two genes suggest that the function of the two genes is slightly different in different organs.

Amino Acid Sequence↗
PubMed2002

Genome sequence and comparative analysis of the model rodent malaria parasite Plasmodium yoelii yoelii.

Species of malaria parasite that infect rodents have long been used as models for malaria disease research. Here we report the whole-genome shotgun sequence of one species, Plasmodium yoelii yoelii, and comparative studies with the genome of the human malaria parasite Plasmodium falciparum clone 3D7. A synteny map of 2,212 P. y. yoelii contiguous DNA sequences (contigs) aligned to 14 P. falciparum chromosomes reveals marked conservation of gene synteny within the body of each chromosome. Of about 5,300 P. falciparum genes, more than 3,300 P. y. yoelii orthologues of predominantly metabolic function were identified. Over 800 copies of a variant antigen gene located in subtelomeric regions were found. This is the first genome sequence of a model eukaryotic parasite, and it provides insight into the use of such systems in the modelling of Plasmodium biology and disease.

Animals↗
PubMed2000

Expression and genome organization of resistance gene analogs in soybean.

Sequence analysis of cloned plant disease-resistance genes reveals a number of conserved domains. Researchers have used these domains to amplify analogous sequences, resistance gene analogs (RGAs), from soybean and other crops. Many of these RGAs map in close proximity to known resistance genes. While this technique is useful in identifying potential disease resistance loci, identifying the functional resistance gene from a cluster of homologs requires sequence information from outside of these conserved domains. To study RGA expression and to determine the extent of their similarity to other plant resistance genes, two soybean cDNA libraries (root and epicotyl) were screened by hybridization with RGA class-specific probes. cDNAs hybridizing to RGA probes were detected in each library. Two types of cDNAs were identified. One type was full-length and contained several disease-resistance gene (R-gene) signatures. The other type contained several deletions within these signatures. Sequence analyses of the cDNA clones placed them in the Toll-Interleukin-1 receptor, nucleotide binding domain, and leucine-rich repeat family of disease-resistance genes. Using clone-specific primers from within the 3' end of the LRRs, we were able to map two cDNA clones (LM6 and MG13) to a BAC contig that is known to span a cluster of disease-resistance genes.

Amino Acid Sequence↗
PubMed2002

The diabetes-prone BB rat carries a frameshift mutation in Ian4, a positional candidate of Iddm1.

Diabetes-prone (DP) BB rats spontaneously develop insulin-dependent diabetes resembling human type 1 diabetes. They also exhibit lifelong T-cell lymphopenia. Functional and genetic data support the hypothesis that the gene responsible for the lymphopenia, Lyp, is also a diabetes susceptibility gene, named Iddm1. We constructed a 550-kb P1-derived artificial chromosome contig of the region. Here, we present a corrected genetic map reducing the genetic interval to 0.2 cM and the physical interval to 150-290 kb. A total of 13 genes and six GenomeScan models are assigned to the homologous human DNA segment on HSA7q36.1, 8 of which belong to the family of immune-associated nucleotides (Ian genes). Two of these are orthologous to mouse Ian1 and -4, both excellent candidates for Iddm1. In normal rats, they are expressed in the thymus and T-cell regions of the spleen. In the thymus of lymphopenic rats, Ian1 exhibits wild-type expression patterns, whereas Ian4 expression is reduced. Mutational screening of their coding sequences revealed a frameshift mutation in Ian4 among lymphopenic rats. The mutation results in a truncated protein in which the COOH-terminal 215 amino acids-including the anchor localizing the protein to the outer mitochondrial membrane-are replaced by 19 other amino acids. We propose that Ian4 is identical to Iddm1.

Amino Acid Sequence↗
PubMed2001

Aberrant hypermethylation of the major breakpoint cluster region in 17p11.2 in medulloblastomas but not supratentorial PNETs.

Deletions of 17p have been consistently reported in up to 50% of medulloblastomas (MBs), and the major breakpoint interval has been localized to chromosome segment 17p11.2. Based on several reports linking aberrant DNA methylation and chromosomal disruption, we examined the methylation pattern in this region by employing restriction landmark genomic scanning (RLGS). Several CpG islands located in the major breakpoint cluster region were identified using a bacterial artificial chromosome (BAC) contig of the breakpoint region. A long-range methylation map was established for 20 MBs and 5 supratentorial primitive neuroectodermal tumors (stPNETs). Selected CpG islands were examined using Southern and bisulfite sequencing analysis. Aberrantly hypermethylated CpG islands in 17p11. 2 were found in 33% of MBs. Interestingly, one CpG island was methylated in MBs, but not in any of the examined stPNETs. A BAC clone covering three of the methylated CpG islands was partially sequenced in the search for a potential tumor suppressor gene. None of the expressed sequence tag sequences and full-length mouse/human cDNAs that were associated with aberrant methylation showed a change in expression levels due to methylation. The potential link between chromosomal instability in 17p11.2 and hypermethylation in this region is discussed.

Adolescent↗
PubMed1998

[Construction and characteristics of a cosmid library of genes of the bacterium Cornyebacterium glutamicum ATSS13032].

A representative genomic library of the Corynebacterium glutamicum ATCC 13032 genes in a cosmid vector Lorist6 was created. The cosmids contain inserts of bacterial DNA obtained by partial digestion with the Sau3A I restrictase. Five hundred and thirty individual primary recombinant clones were transferred into the wells of microtiter plates, where they are now being preserved. The average size of the bacterial DNA inserts determined via a sum of restriction fragment sizes of recombinant molecules is about 38 kb. The capacity of the obtained gene library is 8.4 equivalents of the C. glutamicum genome, i.e., every fragment of the genome is on average represented by eight clones and is presented in at least one clone with the probability > 99%. Clone grids (sets of recombinant clones located on the hybridization membrane in regular and reproducible order) were created. Specificity of the created clone library and its representativeness were confirmed experimentally by hybridization of clone grids with DNA probes corresponding to unique regions of the Corynebacterium genome. A plasmid containing the pheA prephenate dehydratase gene, olygonucleotide corresponding to the lysC gene, and the 21 RNA probe obtained from the insert ends in different cosmids were used as probes. The created set of clones allows the construction of a cosmid contig overlapping the C. glutamicum genome and a physical genetic map on its base.

Corynebacterium↗
PubMed1997

Molecular delineation of the smallest commonly deleted region of chromosome 5 in malignant myeloid diseases to 1-1.5 Mb and preparation of a PAC-based physical map.

Loss of a whole chromosome 5 or a deletion of the long arm, del(5q), is a recurring abnormality in malignant myeloid diseases. In previous studies, we delineated a commonly deleted segment of approximately 4 Mb within band 5q31 that was flanked by IL9 on the proximal side and D5S166 on the distal side. We have generated a physical map of P1 (PAC), bacterial (BAC), and yeast artificial chromosome (YAC) clones of this interval. The contig consists of 108 clones (78 PACs, 2 BACs, and 28 YACs) to which 125 markers (5 genes, 11 expressed sequence tags, 12 polymorphisms, and 97 sequence-tagged sites) have been mapped. Using PAC clones for fluorescence in situ hybridization analysis of leukemia cells with a del(5q), we have narrowed the commonly deleted segment to 1-1.5 Mb between D5S479 and D5S500. To search for allele loss, we used 7 microsatellite markers within and flanking the commonly deleted segment to examine leukemia cells from 28 patients with loss of 5q, and 14 patients without cytogenetically detectable loss of 5q. In the first group of patients, we detected hemizygous deletions, consistent with the cytogenetically visible loss; no homozygous deletions were detected. No allele loss was detected in patients without abnormalities of chromosome 5, suggesting that allele loss on 5q is the result of visible chromosomal abnormalities. The development of a stable PAC contig and the identification of the smallest commonly deleted segment will facilitate the molecular cloning of a myeloid leukemia suppressor gene on 5q.

Chromosome Banding↗
PubMed1993

Theoretical predictions and experimental observations of genomic mapping by anchoring random clones.

Genome mapping by anchoring random clones has recently been the subject of intensive theoretical study. In this paper, differences between published predictions of properties of anchored groups of clones ("contigs") are analyzed and simplifications of the mathematical formulae describing these properties are presented. The theoretical predictions are compared with the experimental results from the physical mapping of the genome of Schizosaccharomyces pombe. Information about the number of genome sections with no anchored clone on them ("oceans") and the number of undetected overlaps between the contigs at a given stage of the experiment is required for the decision to change from the random strategy to that of a directed closure of gaps. We demonstrate that the expected number of oceans can be approximated by the number of groups of clones anchored by a single probe ("singletons"), as can the expected number of undetected overlaps between contigs by the number of contigs containing more than one anchor.

Chromosome Mapping↗
PubMed2004

Phytophthora sojae avirulence genes Avr4 and Avr6 are located in a 24kb, recombination-rich region of genomic DNA.

A cross between two different races (race 7xrace 25) of the soybean root and stem rot pathogen Phytophthora sojae was analyzed to characterize the genomic region flanking two cosegregating avirulence genes, Avr4 and Avr6. Both genes cosegregated in the ratio of 82:17 (avirulent:virulent) in an F(2) population, suggestive of a single locus controlling both phenotypes. A chromosome walk was commenced from RAPD marker OPE7.1C, 2.0cM distant from the Avr4/6 locus. Three overlapping cosmids were isolated which included genetic markers that flank the Avr4/6 locus. The chromosome walk spanned a physical distance of 67kb which represented a genetic map distance of 22.3cM, an average recombination frequency of 3.0kb/cM and 11.7-fold greater than the predicted average recombination frequency of 35.3kb/cM for the entire P. sojae genome. Six genes (cDNA clones) expressed from the Avr4/6 genomic region encompassed by the cosmid contig were identified. Single nucleotide polymorphisms and restriction fragment length polymorphisms showed these six genes were closely linked to the Avr4/6 locus. Physical mapping of the cDNA clones within the cosmid contig made it possible to deduce the precise linkage order of the cDNAs. None of the six cDNA clones appear to be candidates for Avr4/6. We conclude that two of these cDNA clones flank a physical region of approximately 24kb and 4.3cM that appears to include the Avr4/6 locus.

Algal Proteins↗
PubMed1995

TEL and KIP1 define the smallest region of deletions on 12p13 in hematopoietic malignancies.

Unbalanced translocations as well as interstitial deletions of the short arm of chromosome 12 [del(12p)] are found as recurring chromosomal changes in a broad spectrum of hematopoietic malignancies. These changes result in the hemizygous deletion of genetic material from 12p. We mapped a yeast artificial chromosome containing the TEL gene, a cosmid contig containing part of TEL and a P1 contig containing the KIP1 gene to 12p13. These probes were used for fluorescence in situ hybridization to analyze samples from 47 patients with various hematologic malignancies who had unbalanced translocations (25 patients) leading to loss of 12p or deletions (22 patients) involving 12p13. The patients had acute lymphoblastic leukemia (8 cases), myelodysplastic syndrome (MDS; 11 cases), acute myeloid leukemia (AML; 10 cases), myeloproliferative disorders (4 cases), therapy-related MDS or AML (7 cases), non-Hodgkin's lymphoma (2 cases), and other hematopoietic malignancies (5 cases). All three probes were hemizygously detected in 26 cases and were completely retained in only 9 cases. In 12 cases probes for one of the two genes were deleted, allowing us to map the smallest region of overlap of these deletions to a small genomic region that is bordered on the telomeric side by the TEL gene and on the centromeric side by KIP1. The genomic distance between TEL and KIP1 is estimated to be about 1 to 2 Mbp.

Adolescent↗
PubMed1995

Yeast artificial chromosome (YAC)-based genome mapping of Schistosoma mansoni.

Schistosoma mansoni has 7 pairs of autosomal chromosomes and one pair of sex chromosomes (ZZ for a male worm and ZW for a female), of a haploid genome size of 2.7 x 10(8) bp. We initiated the molecular genetic approach for the detailed characterization and understanding of the evolutionary biology of schistosomes. We have constructed a yeast artificial chromosome (YAC)-library with partially digested parasite genomic DNA, and the chromosome location of each insert was detected by fluorescent in situ hybridization (FISH). The library contains > 2283 clones with an average insert size of 358 kb, which represents a 2.6-fold coverage of the genome (> 7.2 x 10(8) bp). 100 randomly selected YAC clones were localized by FISH and found to be distributed widely among all chromosomes. The assembly of 14 YACs distributed almost the whole region of chromosome 3. Generated expressed sequenced tags (ESTs) derived from a unidirectional cDNA library were also used for further characterization of the YAC inserts. These results indicate that an extensive contig assembly of the entire chromosomes and a reasonably detailed gene map should be feasible in the near future.

Animals↗
PubMed1997

Construction of a 780-kb PAC, BAC, and cosmid contig encompassing the minimal critical deletion involved in B cell chronic lymphocytic leukemia at 13q14.3.

A putative tumor suppressor gene involved in B cell chronic lymphocytic leukemia (B-CLL) was mapped to human chromosome 13q14.3 close to the genetic markers D13S25 and D13S319. We constructed a 780-kb-long contig composed of cosmids, bacterial artificial chromosomes, and bacteriophage P1-derived artificial chromosomes that provides essential information and tools for the positional cloning of this gene. The conting contains both flanking markers as well as several additional genetic markers, three ESTs, and one potential CpG island. In addition, using one B-CLL patient, we characterized a small internal deleted region of 550 kb. Comparing this deletion with other recently published deletions narrows the minimally deleted area to less than 100 kb in our physical map. This deletion core region should contain all or part of the disrupted in B cell malignancies tumor suppressor gene.

Bacteriophages↗