Semiautomated DNA probe mapping using digital imaging microscopy: II. System performance.

This paper describes an evaluation of a semiautomated, multicolor image-analysis system to map cloned probes along metaphase chromosomes. Mapping with this system consists of fluorescence in situ hybridization (FISH) for probe localization, automatic acquisition of multicolor images showing total chromosomal DNA and probe location(s), and automatic determination of the fractional locations of the probes along the chromosomes relative to the short arm telomere (FLpter). The system was evaluated by mapping ten phage and ten cosmid probes previously mapped to chromosome 3 with other procedures. The standard deviations of FLpter measurements averaged 3.4 Mb and 2.6 Mb for phage and cosmid probes, respectively. With this variation, the order of two probes mapped in separate hybridizations could be determined with 95% confidence when their separation was greater than 2.5 Mb. In all cases, the probe locations and order were consistent with previous mapping data. FLpter values were converted to band locations using measurements of the band locations made using digital imaging microscopy. This proved superior to conversions made using ISCN ideograms.

[1]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[2]  H. Magdelenat,et al.  Mapping of single-copy DNA sequences on human chromosomes by in situ hybridization with biotinylated probes: enhancement of detection sensitivity by intensified-fluorescence digital-imaging microscopy. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Ashburner,et al.  PCR amplification of DNA microdissected from a single polytene chromosome band: a comparison with conventional microcloning. , 1989, Nucleic acids research.

[4]  D. Moore,et al.  How large should my study be so that I can detect an altered sex ratio? , 1988, Fertility and sterility.

[5]  D. Ward,et al.  Simultaneous visualization of seven different DNA probes by in situ hybridization using combinatorial fluorescence and digital imaging microscopy. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[6]  Iscn.  International System for Human Cytogenetic Nomenclature , 1978 .

[7]  G. Lathrop,et al.  Simultaneous localization of cosmids and chromosome R-banding by fluorescence microscopy: application to regional mapping of human chromosome 11. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[8]  L. S. Cram,et al.  A highly conserved repetitive DNA sequence, (TTAGGG)n, present at the telomeres of human chromosomes. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[9]  B. Trask,et al.  Chromosome heteromorphism quantified by high-resolution bivariate flow karyotyping. , 1989, American journal of human genetics.

[10]  J. Yunis.  High resolution of human chromosomes. , 1976, Science.

[11]  D. Engelke,et al.  Protein-DNA interactions in vivo--examining genes in Saccharomyces cerevisiae and Drosophila melanogaster by chromatin footprinting. , 1991, Methods in cell biology.

[12]  B. Trask,et al.  Mapping of human chromosome Xq28 by two-color fluorescence in situ hybridization of DNA sequences to interphase cell nuclei. , 1991, American journal of human genetics.

[13]  H. Drabkin,et al.  Development of a somatic cell hybrid mapping panel and molecular probes for human chromosome 3. , 1990, Genomics.

[14]  J Vrolijk,et al.  Fluorescence ratio measurements of double-labeled probes for multiple in situ hybridization by digital imaging microscopy. , 1992, Cytometry.

[15]  J. Mcneil,et al.  Interphase and metaphase resolution of different distances within the human dystrophin gene. , 1990, Science.

[16]  N. Tommerup,et al.  A 2.5-Mb physical map within 3p21.1 spans the breakpoint associated with Greig cephalopolysyndactyly syndrome. , 1991, Genomics.

[17]  T. Shows,et al.  Mapping small DNA sequences by fluorescence in situ hybridization directly on banded metaphase chromosomes. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[18]  U. Francke.  Digitized and differentially shaded human chromosome ideograms for genomic applications. , 1994, Cytogenetics and cell genetics.

[19]  G Hermanson,et al.  High-resolution mapping of human chromosome 11 by in situ hybridization with cosmid clones. , 1990, Science.

[20]  J. Piper,et al.  Automatic fluorescence metaphase finder speeds translocation scoring in FISH painted chromosomes. , 1994, Cytometry.

[21]  B. Trask,et al.  Fluorescence in situ hybridization with DNA probes. , 1990, Methods in cell biology.

[22]  D. Ward,et al.  In situ hybridization banding of human chromosomes with Alu-PCR products: a simultaneous karyotype for gene mapping studies. , 1991, Genomics.