Evonetix Limited

Daniel Adrian Bygrave

Matthew James Hayes

Joseph Brennan

Raquel Maria Sanches-Kuiper

Sangeeta Aditya

Raquel Sanches-kuiper

A method for identifying a group of single-stranded oligonucleotides for self-assembly into a double-stranded polynucleotide, the group comprising a plurality of overlapping complementary oligonucleotides, wherein each overlap between complementary oligonucleotides is selected to have a melting temperature (Tm) that differs from the melting temperatures of all other overlapping complementary oligonucleotides in the group.

Method for producing double stranded polynucleotides based on oligonucleotides with selected and different melting temperatures

A series of hybridisations is performed for forming a target double-stranded nucleic acid from initial fragments, where each further hybridisation step hybridises the direct products of a pair of earlier hybridisation steps. For at least one further hybridisation step H, both of the corresponding pair of earlier hybridisation steps Hcomprise an error-detecting type of hybridisation step, which includes an error detecting operation to detect whether the hybridised fragments formed in the error-detecting type of hybridisation step Hcomprise at least one erroneous hybridised fragment, and discarding at least part of the erroneous fragment to exclude it from a subsequent further hybridisation step. By detecting and removing erroneous fragments throughout a staged and controlled hybridisation process, erroneous fragments are prevented from diluting the pool of error-free fragments at each hybridisation step, to improve yield.

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Cambridge, United Kingdom

Daniel Adrian Bygrave