Molecular cloning and characterization of a 20q13.2 amplicon in breast carcinoma
- Lawrence Livermore National Lab., CA (United States); and others
Comparative genomic hybridization (CGH) has identified an amplification event involving chromosome band 20q13.2 in 15-20% of primary breast carcinomas. The application of FISH to the study of tumor interphase nuclei using 33 locus specific cosmid and P1 probes revealed amplification of band 20q13.2 in 35% of breast cancer cell lines and 8% of primary tumors. Moreover, this study localized the amplification event to the 1.5Mb interval defined by (Flpter 0.80-0.84.) and excluded all known genes in the region as candidates for the putative oncogene(s). To both identify the putative oncogene(s) and characterize the amplicon, a 12 member 4Mb YAC contig has been assembled by STS mapping that spans the core of the amplicon. The YAC contig is now being converted to a P1 contig to facilitate sequencing, exon trapping and direct selection of cDNAs. This is being accomplished by performing interAlu PCR reactions on individual YACs and sequencing the reaction products to create 5-10 new STSs per megaYAC. The DuPont P1 library is then screened for these STSs by the PCR. To date 21 P1 clones, forming 6 contigs, have been isolated by screening the DuPont P1 library for existing and/or newly created STSs. The ends of the 21 P1 clones are being sequenced to facilitate contig alignment and to enable chromosome walking. In collaboration with the Human Genome Center at the Lawrence Berkeley Laboratory we have initiated the directed sequencing of two P1 contigs, localized within the amplicon core, and ultimately will sequence the entire 1-2Mb amplicon.
- DOE Contract Number:
- AC03-76SF00098; W-7405-ENG-48; W-7405-ENG-36
- OSTI ID:
- 133485
- Report Number(s):
- CONF-941009--; CNN: Grant 58207
- Journal Information:
- American Journal of Human Genetics, Journal Name: American Journal of Human Genetics Journal Issue: Suppl.3 Vol. 55; ISSN 0002-9297; ISSN AJHGAG
- Country of Publication:
- United States
- Language:
- English
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