Abstract: Disclosed herein are mutants of bacteriophage Phi29 DNA polymerase with improved primer recognition, compared to the wild-type enzyme. Certain mutants comprise one or both of the mutations K64R or M97K. The provided mutants are capable of using more efficiently shorter and longer random synthetic DNA primers than wild-type Phi29 DNA polymerase, generating more amplification product in Multiple Displacement Amplification (MDA) reactions. The inventive mutants amplify human genomic DNA with less bias and better coverage in comparison to reactions carried out with wild-type Phi29 DNA polymerase.

Abstract: Disclosed herein are devices and methods for amplifying double stranded DNA molecules. Methods for amplifying apoptotic cell-free DNA molecules can include performing end-repair and dA tailing of the cfDNA molecules, attachment of single-stranded hairpin adaptors to both ends of the end-repaired cfDNA molecules to produce adaptor-tagged, single-stranded, covalently closed DNA molecules, and amplification of the adaptor-tagged, single-stranded, covalently closed DNA molecules by a combination of rolling circle amplification and multiple displacement amplification using a PrimPol enzyme, a DNA polymerase with strand displacement activity and free nucleotides.

Abstract: The present invention is directed to methods for replicating, amplifying, and sequencing of nucleic acids using the thermostable, bifunctional replicase “TthPrimPol” from Thermus thermophilus HB27. The TthPrimPol enzyme is extremely tolerant to alterations of the nucleotides of the template nucleic acid. Therefore, in one aspect the invention discloses methods for replicating, amplifying and sequencing of damaged polynucleotide templates.

Abstract: Disclosed herein are devices and methods for amplifying double stranded DNA molecules. Methods for amplifying apoptotic cell-free DNA molecules can include performing end-repair and dA tailing of the cfDNA molecules, attachment of single-stranded hairpin adaptors to both ends of the end-repaired cfDNA molecules to produce adaptor-tagged, single-stranded, covalently closed DNA molecules, and amplification of the adaptor-tagged, single-stranded, covalently closed DNA molecules by a combination of rolling circle amplification and multiple displacement amplification using a PrimPol enzyme, a DNA polymerase with strand displacement activity and free nucleotides.