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R H Reeves

Publications and source records attributed to R H Reeves.

At least 37 records · Page 2Linked to original sources
PubMed1999

High-resolution comparative physical mapping of mouse chromosome 10 in the region of homology with human chromosome 21.

Comparative mapping of human and mouse chromosomes can be used to predict locations of homologous loci between the species, provides the substrate to examine the process of chromosomal evolution, and facilitates the continuing development of mouse genetic models for human disorders. A YAC contig of the region of mouse Chromosome (Chr) 10 (MMU10) that demonstrates conserved linkage with the distal portion of human Chr 21 (HSA21) has been constructed. The contig contains all known genes mapped in both species, defines the proximal region of homology between MMU10 and HSA22, and contains the evolutionary junction between HSA21 and HSA22 on MMU10. It consists of 23 YACs and 2 PACs, and covers 3.2 Mb of MMU10. The average marker density for this region is 1 marker/69 kb. Nine of 22 expressed sequences are mapped here for the first time in mouse, and two are newly characterized expressed sequences. The contig also contains 12 simple sequence repeats (SSRs) and 16 YAC and PAC endclone markers. YAC fragmentation analysis was used to create a physical map for the proximal 2.2 Mb of the contig. Cloning of the corresponding region of HSA21 has proven difficult, and the mouse contig includes segments absent from previously described sequence ready maps of HSA21.

Animals↗
PubMed1999

Sequence-ready physical map of the mouse chromosome 16 region with conserved synteny to the human velocardiofacial syndrome region on 22q11.2.

Proximal mouse Chromosome (Chr) 16 shows conserved synteny with human Chrs 16, 8, 22, and 3. The mouse Chr 16/human Chr 22 conserved synteny region includes the DiGeorge/Velocardiofacial syndrome region of human Chr 22q11.2. A physical map of the entire mouse Chr 16/human Chr 22 region of conserved synteny has been constructed to provide a substrate for gene discovery, genomic sequencing, and animal model development. A YAC contig was constructed that extends ca. 5.4 Mb from a region of conserved synteny with human Chr 8 at Prkdc through the region conserved with human Chr 3 at DVL3. Sixty-one markers including 37 genes are mapped with average marker spacing of 90 kb. Physical distance was determined across the 2.6-Mb region from D16Mit74 to Hira with YAC fragmentation. The central region from D16Jhu28 to Igl-C1 was converted into BAC and PAC clones, further refining the physical map and providing sequence-ready template. The gene content and borders of three blocks of conserved linkage between human Chr 22q11.2 mouse Chr 16 are refined.

Animals↗
PubMed1999

Perfect conserved linkage across the entire mouse chromosome 10 region homologous to human chromosome 21.

The distal end of human Chromosome (HSA) 21 from PDXK to the telomere shows perfect conserved linkage with mouse Chromosome (MMU) 10. This region is bounded on the proximal side by a segment of homology to HSA22q11.2, and on the distal side by a region of homology with HSA19p13.1. A high-resolution PAC-based physical map is described that spans 2.8 Mb, including the entire 2.1 Mb from Pdxk to Prmt2 corresponding to HSA21. Thirty-four expressed sequences are mapped, three of which were not mapped previously in any species and nine more that are mapped in mouse for the first time. These genes confirm and extend the conserved linkage between MMU10 and HSA21. The ordered PACs and dense STS map provide a clone resource for biological experiments, for rapid and accurate mapping, and for genomic sequencing. The new genes identified here may be involved in Down syndrome (DS) or in several genetic diseases that map to this conserved region of HSA21.

Animals↗
PubMed1998

A cluster of keratin-associated proteins on mouse chromosome 10 in the region of conserved linkage with human chromosome 21.

A gene cluster of three to five high-cysteine keratin-associated proteins (KAPs) has been identified on mouse Chromosome 10 (MMU10) in the region of conserved linkage with human chromosome 21 (HSA21). One of these genes, Krtap12-1, has been sequenced in its entirety and shown to be an intronless gene encoding a predicted 130-amino-acid protein. Krtap12-1 is most closely related to two previously identified KAP4 genes, but variation in sequence and cysteine content suggests that it represents a new KAP family. Krtap12-1 is expressed in the skin of a 3-day-old mouse. The corresponding region of HSA21, between ITGB2 (integrin beta2) and PFKL (the liver isoform of phosphofructokinase), has proven refractory to cloning, and thus mapping of this region at high resolution has been problematic. Based on the KAP gene cluster position in mouse, evidence has been found for an orthologous human KAP cluster on HSA21q22.3, reinforcing the observation that comparative genomics can play an essential and practical role in determining mammalian genome organization.

Amino Acid Sequence↗
PubMed1998

Localization of tumor suppressor activity important in nonsmall cell lung carcinoma on chromosome 11q.

Loss of heterozygosity on chromosome 11q23 is observed at high frequency in human nonsmall cell lung carcinomas (NSCLCs), suggesting the presence of a tumor suppressor gene. Previous analysis of DNA from 79 patients identified a commonly deleted segment of 5 centimorgans. Complementation analysis was used to further localize a putative tumor suppressor gene. Three yeast artificial chromosome (YAC) clones spanning the minimal loss of heterozygosity region were modified, and spheroplast fusion was used to transfer them into human A549 NSCLC or murine Lewis lung carcinoma (LLC) cell lines. The resulting yeast x human hybrid cell lines containing an intact copy of a 1.6-Mb YAC, 939b12, showed reduced growth in vitro. Injection of parental A549 cells into athymic (nu/nu) mice resulted in tumor formation at 27 of 28 injection sites. In contrast, two independent 939b12-containing cell lines formed tumors at only 3 of 20 injection sites. 939b12 also suppressed tumor formation by LLC NSCLC cells in nude mice, but YACs 785e12 and 911f2, which flank 939b12, had no suppressor activity. Further localization of tumor suppression activity on 939b12 was accomplished by introduction of defined fragmentation derivatives into A549 cells and by analysis of YACs that were broken on transfer into LLC cells. This complementation approach localized tumor suppression activity to the central 700 kb of 939b12 and provides a functional assay for positional cloning of this tumor suppressor gene.

Animals↗
PubMed1998

Physical mapping of the evolutionary boundary between human chromosomes 21 and 22 on mouse chromosome 10.

Adjacent regions of mouse Chromosome 10 (MMU10) show conserved synteny with human chromosome 22 (HSA22) and the telomeric region of HSA21. Physical mapping on MMU10 using YAC fragmentation and PAC contig analyses demonstrates that Prmt2 has a position consistent with its human homolog, HRMT1L1, being telomeric to S100B on HSA21. This result establishes Prmt2 as the new proximal boundary of the region of conserved synteny between MMU10 and HSA21 and predicts that it is the most telomeric gene known on HSA21. Physical mapping refines the positions and order of HSA22 homologs Mmp11, Mif, and Ddt, demonstrates the orientation of S100b on the mouse chromosome, and localizes the junction of conserved synteny between HSA21 and HSA22 on MMU10. Comparative mapping in this region is important for defining gene structure and dosage imbalance in Down syndrome (DS), for developing animal models of DS, and for understanding processes of chromosome evolution.

Animals↗
PubMed1998

Solid-state DNA sizing by atomic force microscopy.

Atomic force microscopy (AFM) allows rapid, accurate, and reproducible visualization of DNA adsorbed onto solid supports. The images reflect the lengths of the DNA molecules in the sample. Here we propose a solid-state DNA sizing (SSDS) method based on AFM as an analytical method for high-throughput applications such as finger-printing, restriction mapping, +/- screening, and genotyping. For this process, the sample is first deposited onto a solid support by adsorption from solution. It is then dried and imaged under ambient conditions by AFM. The resulting images are subjected to automated determination of the lengths of the DNA molecules on the surface. The result is a histogram of sizes that is similar to densitometric scans of DNA samples separated on gels. A direct comparison of SSDS with agarose gel electrophoresis for +/- screening shows that it produces equivalent results. Advantages of SSDS include reduced sample size (i.e., lower reagent costs), rapid analysis of single samples, and potential for full automation using available technology. The high sensitivity of the method also allows the number of polymerase chain reaction cycles to be reduced to 15 or less. Because the high signal-to-noise ratio of the AFM allows for direct visualization of DNA-binding proteins, different DNA conformations, restriction enzymes, and other DNA modifications, there is potential for dramatically improving the information content in this type of analysis.

Adsorption↗
PubMed1998

Exploring development and disease through germ-line genetic engineering in the mouse.

New developments in mouse genetics have transformed the field of genetics from a reactive to a proactive science. Current technologies in transgenesis, chromosome engineering, and saturation mutagenesis are introduced. These approaches permit the creation of new mutations and consequent mutant phenotypes that facilitate both the understanding of functions of existing genes and the search for previously unidentified genes affecting important phenotypes in all mammals.

Animals↗
PubMed1998

Automated sizing of DNA fragments in atomic force microscope images.

Current techniques used to measure lengths of DNA fragments in atomic force microscope (AFM) images require a user to operate interactive software and execute tedious error-prone cursor selections. An algorithm is proposed which provides an automated method for determining DNA fragment lengths from AFM images without interaction from the computer operator (e.g. cursor selections or mouse clicks). The approach utilises image processing techniques tailored to characteristics of AFM images of DNA fragments. The automated measurements have a mean absolute deviation of less than 1 pixel when compared to manual image-based measurements. The DNA length determined from the histogram of calculated lengths is accurate to within 3% of the actual DNA length in solution. For fragments that are 250 base-pairs long, the precision is estimated to be within 17 nm, which is about 20% of the total length. This precision was confirmed when the algorithm easily resolved fragments in one image that differed by only 17 nm. Fragment sizes up to 2000 base-pairs have been tested and successfully sized. This algorithm is being developed as part of a new solid-state DNA sizing technique for applications such as genotyping and construction of physical genome maps.

Algorithms↗
PubMed1998

Genetic component in the inflammatory response induced by bacterial lipopolysaccharide.

Multiple organ dysfunction syndrome (MODS) appears to be the result of a complex program influenced by multiple factors, including environmental, physiological, and immunological conditions. Thus, an uncontrolled inflammatory response following a stochastic event, the initial injury, is believed to be the cause for the development of this syndrome. Several lines of evidence suggest that a genetic component could contribute to the regulation of the inflammatory response, as well, but no direct evidence demonstrates a heritable predisposition to MODS. In the present study, a genetic contribution was demonstrated for the inflammatory response induced by the administration of bacterial lipopolysaccharide (LPS) in different, genetically distinct strains of inbred mice. A survey of five inbred strains showed that mortality following administration of Escherichia coli LPS (20 mg/kg) was highest in C57BL/6J (B6) mice, while A/J mice were the most resistant. Accordingly, B6 and A/J mice were examined further for differences in the inflammatory response elicited by LPS. B6 mice showed higher levels of circulating interleukin-1beta and interleukin-6, as well as higher mRNA levels of hepatic beta-fibrinogen (an acute-phase gene) and metallothionein. Surprisingly, the circulating levels of tumor necrosis factor-alpha were significantly higher in A/J than in B6 mice after LPS administration. Since B6 and A/J mice were bred and raised in identical environments and received the same LPS challenge, the contrasting inflammatory response that was observed is largely attributable to genetic differences between these two strains. These data illustrate that the response to injury could be modulated by the genetic background of the individual. This information may be pertinent for the care of critically ill patients.

Animals↗
PubMed1998

Physical and comparative mapping of distal mouse chromosome 16. 5 p5.

Distal mouse Chromosome 16 (Chr. 16) includes a region of conserved linkage with human Chromosome 21 (Chr. 21). Mouse models of Down syndrome based on trisomy of distal Chr. 16 have several phenotypes similar to those seen in human patients and have proven useful for correlating dosage imbalance of specific genes with specific developmental anomalies. The degree to which such findings can be related to Down syndrome depends on how well the conserved synteny is maintained. Twenty-four genes have been mapped in both species and there are no discordancies, but the region could carry hundreds of genes. Comparative sequence represents the ultimate comparative map and will aid in identification of genes and their regulatory sequences. A physical map of the distal 4.5 Mb of Chr. 16 has been assembled as an essential step toward a map of sequence-ready templates. The map consists of 51 YACs and 15 BACs and includes 18 transcripts, 9 of which are mapped for the first time in mouse, and 3 of which are, for the first time, described in either species. YAC fragmentation was used to precisely localize the 49 markers on the map. Comparison of this physical map with that of the corresponding region on Chr. 21 shows conservation not only of gene order but of size in the 3 Mb from Cbr1 to Ets2; distal to Ets2, the human map is expanded.

Animals↗
PubMed1997

High-resolution recombinational map of mouse chromosome 16.

Five intersubspecific backcrosses and an intercross were used to establish a sex-averaged recombinational map spanning 56 cM across most of mouse Chromosome 16 (Chr 16). A total of 123 markers were ordered using an interval mapping approach to identify 425 recombination sites in a collection of 1154 meioses from 1155 progeny generated in the six crosses. The markers include the 10 "classic" Chr 16 reference markers, 26 additional genes or transcripts including two phenotypic markers (Pit1dw and Kcnj6wv), and 87 simple sequence length polymorphisms (SSLPs). One set of monozygotic twins was detected among the 304 meioses mapped to highest resolution. The reference markers and SSLPs allow the map to be well integrated with existing maps of Chr 16. The average distance between crossover sites is less than 500 kb for most chromosomes, making this collection of recombinant chromosomes useful as a binning and ordering resource for YAC-based physical map assembly on Chr 16.

Animals↗