Abstract We describe an integrated approach to the de- termination of complex Y chromosome haplotypes that is both fast and relatively inexpensive. The method employs GeneScan technology to enable a researcher to assay re- peat number variation at ten microsatellite loci and deter- mine the status of 11 diallelic polymorphisms. The method requires only four PCRs and four GeneScan runs per sam- ple and is relatively insensitive to sample DNA concen- tration. Introduction In recent years the determination of human Y chromo- some haplotypes has received attention from a number of fields including anthropology (Hammer et al. 1998) and forensics (Kayser et al. 1997). Increasing use has been made of compound haplotypes, comprised of microsatel- lites and diallelic polymorphisms, to differentiate popula- tions (Roewer et al. 1996) and to study human migration in prehistory (Zerjal et al. 1997). Thomas et al. (1998) have suggested that a compound Y chromosome haplo- type of six microsatellites and six diallelic, presumed unique event polymorphisms (UEP) may be a “signature haplotype” of the ancient Hebrew population. Microsatel- lite variation has been used to date events in prehistory, al- beit with wide confidence intervals (Kittles et al. 1998; Thomas et al. 1998). All of the studies mentioned require the generation of considerable quantities of data on the al- lelic state of microsatellites and diallelic polymorphisms, a process that can be both expensive and slow. Methods that reduce cost and increase the rate at which data are generated are therefore beneficial. Here, we describe a method for typing 11 previously reported Y chromosome diallelic polymorphisms and 10 commonly used Y chromosome microsatellites. The 11 diallelic polymorphisms define 13 lineages, 11 of which are found at high frequency in at least one major African or Eurasian population. The method requires only four multiplex PCRs and four runs per sample on an ABI-310 genetic analyser or ABI-377 automated sequencer. The multiplex PCR “kits” that permit this high throughput analysis of Y chromosome polymorphisms contain oligo- nucleotide primers labelled with three different ABI dyes (HEX, TET and FAM). The resulting PCR products are analysed in the presence of GS-500 or GS-350 molecular weight standards labelled with the fluorescent dye TAMRA (PE-Applied Biosystems). The amplified DNA fragments generated by the kits are all under 430 base pairs (bp) in length, and most are smaller than 230 bp. It is therefore possible to use the kits to amplify degraded DNA samples, including DNA extracted from blood archives, forensic material and archaeological remains. Microsatellite kit 1 (MS1) contains four previously re- ported primer pairs that amplify DYS19, DYS388, DYS390 and DYS393 (Kayser et al. 1997) and two primer pairs that were redesigned to amplify DYS391 and DYS392 (Table 1). Primers for loci DYS391 and DYS392 were redesigned to ensure that all PCR products from this kit fall within a limited size range (100–230 bp) and that the six microsatellites can be discriminated using a com- bination of size and fluorescent dye label. Microsatellite kit 2 (MS2) contains primer pairs that amplify loci DYS388, DYS389 I, DYS389 II, DYS425 and DYS426 (Jobling et al. 1996; Kayser et al. 1997). To ensure com- patibility under multiplex conditions, it was necessary to redesign five out of the eight primers (Table 2). The DYS388 locus is amplified by both MS1 and MS2 kits, providing a degree of internal control. Two additional multiplex PCR kits were designed to type diallelic polymorphisms at 11 separate loci. Selection of polymorphisms for inclusion in these kits was based on (1) potential usefulness in distinguishing populations and (2) mutual compatibility of markers within the same mul- Mark G. Thomas · Neil Bradman · Helen M. Flinn High throughput analysis of 10 microsatellite and 11 diallelic polymorphisms on the human Y-chromosome Hum Genet (1999) 105 : 577–581 © Springer-Verlag 1999 Digital Object Identifier (DOI) 10.1007/s004399900181 Received: 12 August 1999 / Accepted: 30 September 1999 / Published online: 4 November 1999 ORIGINAL INVESTIGATION M.G. Thomas (✉) · N. Bradman · H.M. Flinn The Centre for Genetic Anthropology, The Departments of Anthropology and Biology, Darwin Building, University College London, Gower Street, London WC1E 6BT, UK e-mail: m.thomas@ucl.ac.uk, Fax: +44-171-3807096