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PLoS One. 2017 Sep 11;12(9):e0184507. doi: 10.1371/journal.pone.0184507. eCollection 2017.

Improved DOP-PCR (iDOP-PCR): A robust and simple WGA method for efficient amplification of low copy number genomic DNA.

Author information

1
All-Russia Institute of Agricultural Biotechnology, Russian Academy of Sciences, Moscow, Russia.
2
Vavilov Institute of General Genetics, Russian Academy of Sciences, Moscow, Russia.
3
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia.
4
Genetic Expertise LLC, Moscow, Russia.
5
Evrogen JSC, Moscow, Russia.
6
Pirogov Russian National Research Medical University, Moscow, Russia.
7
Syntol JSC, Moscow, Russia.
8
Bioline Ltd, London, United Kingdom.
9
Birkbeck, University of London, London, United Kingdom.
10
The Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden.

Abstract

Whole-genome amplification (WGA) techniques are used for non-specific amplification of low-copy number DNA, and especially for single-cell genome and transcriptome amplification. There are a number of WGA methods that have been developed over the years. One example is degenerate oligonucleotide-primed PCR (DOP-PCR), which is a very simple, fast and inexpensive WGA technique. Although DOP-PCR has been regarded as one of the pioneering methods for WGA, it only provides low genome coverage and a high allele dropout rate when compared to more modern techniques. Here we describe an improved DOP-PCR (iDOP-PCR). We have modified the classic DOP-PCR by using a new thermostable DNA polymerase (SD polymerase) with a strong strand-displacement activity and by adjustments in primers design. We compared iDOP-PCR, classic DOP-PCR and the well-established PicoPlex technique for whole genome amplification of both high- and low-copy number human genomic DNA. The amplified DNA libraries were evaluated by analysis of short tandem repeat genotypes and NGS data. In summary, iDOP-PCR provided a better quality of the amplified DNA libraries compared to the other WGA methods tested, especially when low amounts of genomic DNA were used as an input material.

PMID:
28892497
PMCID:
PMC5593185
DOI:
10.1371/journal.pone.0184507
[Indexed for MEDLINE]
Free PMC Article

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