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The CEPH (Centre d'Etude du Polymorphisme Humain) B library (Research Genetics Huntsville AL) was screened by PCR using a series of tiered pools to identify unique clones.

The forward PCR primer was labeled with [ 32 P]ATP and used to amplify 40 ng of genomic DNA. The samples were then subjected to electrophoresis and autoradiography. The samples included 25 human breast tumor cell lines available from American Type Culture Collection (ATCC) as well as 40 human primary breast tumors xenografted into nude mice. The cell lines were HS578T SK-BR-3 UACC812 UACC893 MDA-MB-453 MDA-MB-175-VII MDA-MB468 MDA-MB-361 MDA-MB-231 MDA-MB-436 MDA-MB-415 MDA-MB-330 MDA-MB-157 MDA-MB-134-VI MDA-MB-435S ZR 75-30 ZR 75-1 BT-549 BT-483 T-47D BT-474 DU4475 CAMA1 MCF7 and BT-20.

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BAC DNA was prepared using the Nucleobond kit (Nest Group Southboro MA). BACs were digested with Not I and subjected to electrophoresis on a field inversion apparatus. BACs A B C and D were 240 200 175 and 120 kb respectively (see Fig. 1B). A Not I site was present 20 kb from one end of BAC D. Twelve new STS sites were generated by sequencing both ends of BACs B C and D and shotgun cloning Eco RI fragments. Plasmid DNA was prepared from the cloned Eco RI fragments. DNA was cycle sequenced with appropriate primers using a [ 33 P]ddNTP cycle sequencing kit (Amersham). STS primers were designed and the relative location of the STSs determined by testing for their presence in the BAC contig. Primer sequences are available upon request.

The glioblastoma lines included U105 U118MG A172 DBTRG-05MG U373MG T-98G U-87MG and U138MG and 34 glioblastoma xenografts. The prostate cancer cell lines tested were DU145 LNCaP NCI H660 and PC-3 and microsatellite analysis revealed that each was unique. With the exception of U105 all lines were obtained from ATCC.

DNA (10 μg) was digested with Eco RI resolved on a 1% agarose gel and transferred to nylon. The JL25 3-kb probe and the 2-kb control probe were randomly labeled and hybridized to the blot consecutively.

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The map panel #2 monochromosome panel was purchased from the National Institute of General Medical Science (NIGMS) Human Mutant Genetic Cell Repository.

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STSs Not-5′ PTPD and ET-1 were amplified from primary glioblastoma DNA and blood DNA and the exonic regions were sequenced.

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We thank B. Vogelstein N. Tonks and E. Marcantonio for their comments and S. Kalachikov and R. Hauptschein for helpful suggestions. R.P. is a James S. McDonnell Scholar. M.H.W. is an American Cancer Society Research Professor and is supported by the Department of the Army (DAMD 17-94-I4247) NCI (5R35 CA39829) Amplicon Corporation and the “1 in 9” breast cancer organization. This work is dedicated to the memory of Richard K. Parsons and Richard P. Sanchez.