Genome coverage and sequence fidelity of phi29 polymerase-based multiple strand displacement whole genome amplification.
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.
Paez
J Guillermo
JG
Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA 02115, USA.
Lin
Ming
M
Beroukhim
Rameen
R
Lee
Jeffrey C
JC
Zhao
Xiaojun
X
Richter
Daniel J
DJ
Gabriel
Stacey
S
Herman
Paula
P
Sasaki
Hidefumi
H
Altshuler
David
D
Li
Cheng
C
Meyerson
Matthew
M
Sellers
William R
WR
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
D004259:DNA-Directed DNA Polymerase