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Methods

Clone-Based Systematic Haplotyping (CSH): A Procedure for Physical Haplotyping of Whole Genomes

¹ Max-Planck-Institute for Molecular Genetics, D-14195 Berlin-Dahlem, Germany ² HAEMA AG, 12689 Berlin, Germany

We present a novel methodology to determine the phase of single-nucleotide polymorphisms (SNPs) on a chromosome, which we term clone-based systematic haplotyping (CSH). The CSH procedure is based on separating the allelic chromosomes of a diploid genome by fosmid/cosmid cloning, and subsequent SNP typing of 96 clone pools, each representing 10% of the genome. The pools are screened by PCR for the sequence of interest, followed by SNP typing on the PCR products using the GOOD assay. We demonstrate that by CSH, the haplotype of SNPs separated by more than 50 kilobases can definitely be assigned. We propose this method as being suitable for constructing maps of ancestral haplotypes, analysis of complex diseases, and for diagnosis of rare defects in which the molecular haplotype is crucial. In addition, by amplifying the initial DNA by many orders of magnitude, the original DNA resource is effectively immortalized, enabling the haplotyping of hundreds of thousands of SNPs per individual.

³ Corresponding authors.
E-MAIL burgtorf{at}molgen.mpg.de; FAX 49-30-84131365.
E-MAIL sauer{at}molgen.mpg.de; FAX 49-30-84131365.

[Supplemental material is available online at www.genome.org. The following individuals kindly provided reagents, samples, or unpublished information as indicated in the paper: R. Pawlik.]

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