Sequence Ready Characterization of the Pericentromeric Region of 19p12

Technical Report · Thu Aug 31 04:00:00 EDT 2006

DOI:https://doi.org/10.2172/890705· OSTI ID:890705

Eichler, Evan E ^[1]

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1. University of Washington

Current mapping and sequencing strategies have been inadequate within the proximal portion of 19p12 due, in part, to the presence of a recently expanded ZNF (zinc-finger) gene family and the presence of large (25-50 kb) inverted beta-satellite repeat structures which bracket this tandemly duplicated gene family. The virtual of absence of classically defined “unique” sequence within the region has hampered efforts to identify and characterize a suitable minimal tiling path of clones which can be used as templates required for finished sequencing of the region. The goal of this proposal is to develop and implement a novel sequence-anchor strategy to generate a contiguous BAC map of the most proximal portion of chromosome 19p12 for the purpose of complete sequence characterization. The target region will be an estimated 4.5 Mb of DNA extending from STS marker D19S450 (the beginning of the ZNF gene cluster) to the centromeric (alpha-satellite) junction of 19p11. The approach will entail 1) pre-selection of 19p12 BAC and cosmid clones (NIH approved library) utilizing both 19p12 -unique and 19p12-SPECIFIC repeat probes (Eichler et al., 1998); 2) the generation of a BAC/cosmid end-sequence map across the region with a density of one marker every 8kb; 3) the development of a second-generation of STS (sequence tagged sites) which will be used to identify and verify clonal overlap at the level of the sequence; 4) incorporation of these sequence-anchored overlapping clones into existing cosmid/BAC restriction maps developed at Livermore National Laboratory; and 5) validation of the organization of this region utilizing high-resolution FISH techniques (extended chromatin analysis) on monochromosomal 19 somatic cell hybrids and parental cell lines of source material. The data generated will be used in the selection of the most parsimonious tiling path of BAC clones to be sequenced as part of the JGI effort on chromosome 19 and should serve as a model for the sequence characterization of other difficult regions of the human genome

• Research Organization: Case Western Reserve University
• Sponsoring Organization: USDOE - Office of Science (SC)
• DOE Contract Number: FG02-99ER62862
• OSTI ID: 890705
• Report Number(s): DOE/FG/62862-1
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
CHROMATIN
CHROMOSOMES
COSMIDS
DNA
GENES
Genome sequencing
chromsome 19
PROBES
SOMATIC CELLS
TARGETS
VALIDATION