Genome amplification approaches: DOP-PCR, MDA, MALBAC


DOP-PCR = degenerate oligonucleotide priming polymerase chain reaction
Method Overview: In the first cycling stage (5-8 cycles), low-temperature annealing and extension occur at many binding sites in the genome and become tagged with the DOP primer. In the second cycling stage (>25 cycles), annealing temperature is raised, increasing priming specificity during amplification of the tagged sequence.
Pros: Degeneracy of the primer and the initial template synthesis conditions allow for uniformity of amplification, allowing for faithful recapitulation of copy number states.
Cons: Unable to cover the majority of the genome, though studies have shown up to 40% of the genome of any single cell can be retrieved when sequenced at sufficient depths.

MDA = multiple displacement amplification
Method Overview: Uses phi29 DNA polymerase, which has strand displacement and high processivity. Single stranded DNA templates are generated in the displacement step, and then reprimed and extended, leading to amplification.
Pros: Processivity allows for retrieval of up to 75% of the genome when sequenced at sufficient depths.
Cons: Non-uniform nature of the amplification reaction distorts the copy number landscape.

MALBAC = multiple annealing and lopping-based amplification cycles
Method Overview: Strand displacement pre-amplification generates amplicons with complementary ends that, in theory, mediate molecule looping to prevent further amplification. Pre-amplification is followed by regular PCR to further amplify amplicons.
Pros: High coverage compared to MDA, significantly reduced allelic drop out rate, and appears to maintain the copy number landscape when sequencing at high sequencing coverage and low resolution.
Cons: Low fidelity of polymerase used in generating the initial templates leads to a high false positive SNP rate.

Source: Baslan, T., & Hicks, J. (2014). Single cell sequencing approaches for complex biological systems. Current Opinion in Genetics & Development, 26(0), 59–65. doi:

So, what do you think ?