Quantitative DNA pooling to increase the efficiency of linkage analysis in autosomal dominant disease |
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Authors: | K F Damji C J Gallione R R Allingham B Slotterbeck A E Guttmacher K A Pasyk J M Vance M A Pericak-Vance M C Speer D A Marchuk |
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Institution: | (1) University of Ottawa Eye Institute, 501 Smyth Rd, Ottawa, Ontario, Canada K1H 8L6 Fax: +1-613-737-8836; e-mail: kdamji@ogh.on.ca, CA;(2) Duke University, Durham, N.C., USA, US;(3) University of Vermont College of Medicine, Burlington, Vt., USA, US;(4) University of Michigan, Ann Arbor, Mich., USA, US |
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Abstract: | DNA pooling is an efficient method to rapidly perform genome-wide linkage scans in autosomal recessive diseases in inbred
populations where affected individuals are likely to be homozygous for alleles near the disease gene locus. We wanted to examine
whether this approach would detect linkage in autosomal dominant (AD) disorders where affected individuals may share one allele
identical by descent at loci tightly linked to the disease. Two large outbred pedigrees in which the AD diseases familial
venous malformation (FVM) and hereditary hemorrhagic telangiectasia (HHT1), linked to 9p and 9q, respectively, were investigated.
Separate pools of DNA from affected (n = 21 for FVM and 17 for HHT1) and unaffected family members (n = 9 FVM and HHT1), and 25 unrelated population controls were established. Polymorphic markers spanning chromosome 9 at approximately
13.5-cM intervals were amplified using standard PCR. Allele quantitation was performed with a fluorimager. Visual inspection
of allele intensities and frequency distributions suggested a shift in frequency of the most common allele in the affecteds
lane when compared to control lanes for markers within 30 cM of the FVM and HHT1 loci. These subjective assessments were confirmed
statistically by testing for the difference between two proportions (one-sided; P≤ 0.05). When using population controls, the true-positive rates for FVM and HHT1 were 5/5 and 2/5 markers, respectively.
False-positive rates for FVM and HHT1 were 3/9 and 2/9, respectively. In both AD diseases investigated, quantitative DNA pooling
detected shifts in allele frequency, thus identifying areas of known linkage in most cases. The utility of this technique
depends on the size of the pedigree, frequency of the disease-associated allele in the population, and the choice of appropriate
controls. Although the false-positive rate appears to be high, this approach still serves to reduce the amount of overall
genotyping by about 60%. DNA pooling merits further investigation as a potential strategy in increasing the efficiency of
genomic linkage scans.
Received: 12 May 1997 / Accepted: 29 October 1997 |
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