title: Genome coverage and sequence fidelity of phi29 polymerase-based multiple strand displacement whole genome amplification.
abstract: Major efforts are underway to systematically define the somatic and germline genetic variations causally associated with disease. Genome-wide genetic analysis of actual clinical samples is, however, limited by the paucity of genomic DNA available. Here we have tested the fidelity and genome representation of phi29 polymerase-based genome amplification (phi29MDA) using direct sequencing and high density oligonucleotide arrays probing >10,000 SNP alleles. Genome representation was comprehensive and estimated to be 99.82% complete, although six regions encompassing a maximum of 5.62 Mb failed to amplify. There was no degradation in the accuracy of SNP genotyping and, in direct sequencing experiments sampling 500,000 bp, the estimated error rate (9.5 x 10(-6)) was the same as in paired unamplified samples. The detection of cancer-associated loss of heterozygosity and copy number changes, including homozygous deletion and gene amplification, were similarly robust. These results suggest that phi29MDA yields high fidelity, near-complete genome representation suitable for high resolution genetic analysis.
authors.affiliation: Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA 02115, USA.
authors.forename: J Guillermo
authors.initials: JG
authors.lastname: Paez
authors.forename: Ming
authors.initials: M
authors.lastname: Lin
authors.forename: Rameen
authors.initials: R
authors.lastname: Beroukhim
authors.forename: Jeffrey C
authors.initials: JC
authors.lastname: Lee
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Zhao
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Richter
authors.forename: Stacey
authors.initials: S
authors.lastname: Gabriel
authors.forename: Paula
authors.initials: P
authors.lastname: Herman
authors.forename: Hidefumi
authors.initials: H
authors.lastname: Sasaki
authors.forename: David
authors.initials: D
authors.lastname: Altshuler
authors.forename: Cheng
authors.initials: C
authors.lastname: Li
authors.forename: Matthew
authors.initials: M
authors.lastname: Meyerson
authors.forename: William R
authors.initials: WR
authors.lastname: Sellers
mesh_terms: D000483:Alleles; D017103:Bacillus Phages / Q000201:enzymology*; D002460:Cell Line; D045744:Cell Line, Tumor; D002872:Chromosome Deletion; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D018628:Gene Dosage; D015894:Genome, Human; D023281:Genomics / Q000379:methods*; D005838:Genotype; D006720:Homozygote; D006801:Humans; D019656:Loss of Heterozygosity / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D004259:DNA-Directed DNA Polymerase