Affiliations: Bioinformatics Group, Nanyang Polytechnic, 180 Ang Mo
Kio Avenue 8, Singapore 569830, Republic of Singapore | Division of Medical Sciences, National Cancer Centre,
11 Hospital Drive, Singapore 169610, Republic of Singapore
Note: [] Corresponding author for experimental validation. E-mail:
[email protected]
Abstract: Whole Genome Amplification (WGA) is an important process to increase
limiting amounts of genomic DNA prior to genomic analyses. Current
amplification methods based on primer extension or strand displacement
principles employ primers of partially or totally random sequence. In this
paper, we present a method using Genetic Algorithms to optimize a single primer
design to be used in a primer extension reaction to achieve unbiased WGA.
Computational simulation and prediction of a suitable primer proposed two
candidates NYP6-1 (ATCTCA) and NYP6-2 (TGAGAT). NYP6-1 amplified to a maximum
length of 2537 base pairs (bp), had genome coverage of approximately 45.62%,
with an average of 493 and variance of 163 amplicons per 1 megabasepairs (Mb).
NYP6-2 amplified to a maximum length of 2926 bp and covered 54.35% of the
genome with an average of 579 and a variance of 191 amplicons per Mb. In
contrast, the original primer used in Degenerate Oligonucleotide-Primed PCR
(DOP-PCR) had coverage of 20.93%, an average of 74 and variance of 188
amplicons per Mb when extended up to a length of 2000 bp. Successful WGA of
miniscule amounts of genomic DNA requires the amplification method used to
resolve issues on efficiency, accurate representation of the whole genome and
ability to degraded DNA. The sequence NYP6-2 discovered using our method can be
confidently used in a primer extension based protocol to perform quantitatively
unbiased WGA.
