TruePrime™ is a novel MDA (multiple displacement amplification)-type DNA amplification method that utilizes a primase activity (TthPrimPol) rather than random hexamers to generate DNA-primers. thPrimPol is a monomeric enzyme (34 kDa) that displays a potent primase activity, preferring dNTPs as substrates unlike conventional primases. This DNA primase activity can be activated by magnesium or manganese ions, having wide sequence specificity for template recognition. In this setup, TthPrimPol synthesizes the DNA primers needed for Phi29 DNA pol, which allows for the exponential amplification of genomic DNA. Key advantages of the TruePrime™ technology for amplification of single cell genomes include complete absence of primer artefacts, superior sensitivity down to the femtogram range, and an easy reaction workflow. Analyses on genomic DNA amplified from single Hek293 cells in comparison with non-amplified DNA and the commercially available MDA methods based on random synthetic primers reveal an excellent genome coverage breadth with little bias (~92% at 20x coverage). Chimera formation due to amplification is seen in ~3% of sequenced read pairs. Copy number variant (CNV) detection with algorithms based on read depth analyses is excellent due to the low bias in genome coverage. Low allelic dropout rates allow excellent retrieval of SNVs. We conclude that TruePrime™ is a superior method for genome amplification that builds on the unique advantages of the Phi29-mediated MDA process but overcomes disadvantages of the random-primed protocols used so far.