Whole-genome amplification: relative efficiencies of the current methods.
Whole genome amplification (WGA) has emerged as a fundamental method for DNA analysis from limited quantities of genomic DNA in forensic analysis and disease gene discovery. Several strategies for WGA have been developed during the past decade, each with variable fidelity, yield and coverage of the amplified genome. In the search for a reliable and robust WGA method for genotyping short tandem repeat (STR) loci and single nucleotide polymorphic (SNP) markers, we initially tested four common methods, viz., degenerate oligonucleotide primed PCR (DOP), primer extension preamplification (PEP), improved-PEP (I-PEP) and multiple displacement amplification (MDA), typing the 13 CODIS tetranucleotide repeat loci. The results showed among all methods, I-PEP and MDA have higher genomic coverage. DOP and PEP produced locus and allelic dropouts. Therefore, we emphasized on the evaluation of I-PEP and MDA protocols, which shows that the two methods have their relative strengths and weaknesses. In general, the product yield of MDA is higher than that of I-PEP. However, the specificity of the I-PEP products appears to be higher, particularly in the analysis of STR loci. In the analysis of SNP markers, some loci amplify better using products obtained from I-PEP whereas some worked better with MDA. Our analyses also demonstrate that blood spots on FTA cards are a more efficient source of DNA for I-PEP as compared to MDA, especially for STR analysis.
Department of Environmental Health, Center for Genome Information, University of Cincinnati Medical Center, Cincinnati, OH 45267-0056, USA.
Guangyun
G
Sun
Ritesh
R
Kaushal
Prodipto
P
Pal
Michael
M
Wolujewicz
Diane
D
Smelser
Hong
H
Cheng
Mei
M
Lu
Ranajit
R
Chakraborty
Li
L
Jin
Ranjan
R
Deka
D022981:Allelic Imbalance; D004526:Efficiency; D015894:Genome, Human; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity; D020080:Tandem Repeat Sequences