Long DOP-PCR of rare archival anthropological samples.

The application of molecular DNA technologies to anthropological questions has meant that rare or archival samples of human remains, including blood, hair, and bone, can now be used as a source of material for genetic analysis. Often, these samples are irreplaceable, and/or yield very small quantities of DNA, so methods for preamplifying as much of the whole genome as possible would greatly enhance their usefulness. DOP-PCR (degenerate oligonucleotide-primed polymerase chain reaction) is an amplification method that uses a degenerate primer and very low initial annealing temperatures to amplify the whole genome. We adapted a published DOP-PCR protocol to long PCR enzyme and amplification conditions. The effectiveness of these modifications was tested by PCR amplification of DOP-PCR products at a mixture of genomic targets including 66 different microsatellites, 11 Alu insertion polymorphisms, and variable-length segments of the human lipoprotein lipase gene ( LPL ). The selected microsatellite markers were chosen to represent every chromosome, with expected product sizes ranging from 150 base pairs to 8000 base pairs in length, while the 22 Alu insertion polymorphisms were selected to reveal biases in the recovery of alleles of different sizes. To determine nucleotide sequence variation, 2 kilobases (kb) of the LPL gene in 30 Mongolian individuals were sequenced. All gene-specific targets from DOP-PCR product template were amplified. No unexpected polymorphisms in the sequence results attributable to the DOP-PCR step were found, and 93% to 95% of Alu genotypes that have been amplified from total genomic DNA were replicated. The incorrect typings were all due to the preferential amplification of the shorter of two possible alleles in individuals heterozygous for an Alu insertion and were all correctly typed on subsequent reamplification of 1 Department of Anthropology, The Pennsylvania State University, University Park, PA 16802. 2 Department of Pathology, Biometry, and Genetics, and Department of Biochemistry and Molecular Biology, Stanley S. Scott Cancer Center, Neuroscience Center of Excellence, Louisiana State University Health Sciences Center, New Orleans, LA. 3 Department of Biology, The Pennsylvania State University, University Park, PA. 4 Population Research Institute, The Pennsylvania State University, University Park, PA. 5 Graduate Program in Genetics, The Pennsylvania State University, University Park, PA. Human Biology , December 2000, v. 72, no. 6, pp. 911-925. Copyright © 2000 Wayne State University Press, Detroit, Michigan 48201-1309