Abstract: A method for template-directed sequencing-by-synthesis of an array of target polynucleotide can include: (a) providing an array of target polynucleotides in a fluidic vessel; (b) contacting the array of polynucleotides with a solution comprising (i) polymerization complex and (ii) reversibly terminating and differently labeled A, C, G, and T/U nucleotides; (c) incorporating one of the differently labeled nucleotides, using the polymerization complex, into a chain complementary to at least one of the array of polynucleotides; (d) binding imaging tags to the differently labeled nucleotides of step (c); (e) imaging and storing the identity and position of the imaging tags of step (d); (f) reversing termination (b)-(e); (g) repeating steps (b)-(e) and assembling a sequence for each of the array of target polynucleotides from the stored identity and position of the imaging tags, optionally as a homogeneous or one pot reaction. Additional methods of sequencing target polynucleotides are described herein.

Abstract: A method for template-directed sequencing-by-synthesis of an array of target polynucleotide can include: (a) providing an array of target polynucleotides in a fluidic vessel; (b) contacting the array of polynucleotides with a solution comprising (i) polymerization complex and (ii) reversibly terminating and differently labeled A,C,G, and T/U nucleotides; (c) incorporating one of the differently labeled nucleotides, using the polymerization complex, into a chain complementary to at least one of the array of polynucleotides; (d) binding imaging tags to the differently labeled nucleotides of step (c); (e) imaging and storing the identity and position of the imaging tags of step (d); (f) reversing termination (b)-(e); (g) repeating steps (b)-(e) and assembling a sequence for each of the array of target polynucleotides from the stored identity and position of the imaging tags, optionally as a homogeneous or one pot reaction. Additional methods of sequencing target polynucleotides are described herein.

Abstract: Methods of detecting, probing, mapping and directed sequencing of target nucleic acids are provided using a guide RNA and a Cas9 protein. Methods for detecting the binding of the guide RNA/Cas9 complex to a target nucleic acid where the guide RNA includes a 3? tail sequence that can hybridize to a probe are provided. Methods for detecting the binding of the guide RNA/Cas9 complex to a target nucleic acid where the complex is physically detected are provided.

Abstract: The present invention relates to a method of sequencing a target polynucleotide by enzymatic and/or chemical means. The sequencing method includes a method for characterizing multiple alleles in a sample, a method for comparing polynucleotide sequence. The method involves analysing molecules as members of an array. Many target polynucleotides or many segments of a single target polynucleotide can be sequenced simultaneously. In a preferred embodiment the method involves analysing individual molecules within an array and base calls are based on the signals from two or more molecules. A method to prevent non-specific signal in sequencing is also provided. The invention is readily automated, both for small-scale and large-scale operation and relevant algorithms and the composition of kits and systems are provided.

Abstract: The present invention relates to a method of sequencing a target polynucleotide by enzymatic and/or chemical means. The sequencing method includes a method for characterizing multiple alleles in a sample, a method for comparing polynucleotide sequence. The method involves analysing molecules as members of an array. Many target polynucleotides or many segments of a single target polynucleotide can be sequenced simultaneously. In a preferred embodiment the method involves analysing individual molecules within an array and base calls are based on the signals from two or more molecules. A method to prevent non-specific signal in sequencing is also provided. The invention is readily automated, both for small-scale and large-scale operation and relevant algorithms and the composition of kits and systems are provided.

Abstract: A method for template-directed sequencing-by-synthesis of an array of target polynucleotide can include: (a) providing an array of target polynucleotides in a fluidic vessel; (b) contacting the array of polynucleotides with a solution comprising (i) polymerization complex and (ii) reversibly terminating and differently labeled A,C,G, and T/U nucleotides; (c) incorporating one of the differently labeled nucleotides, using the polymerization complex, into a chain complementary to at least one of the array of polynucleotides; (d) binding imaging tags to the differently labeled nucleotides of step (c); (e) imaging and storing the identity and position of the imaging tags of step (d); (f) reversing termination (b)-(e); (g) repeating steps (b)-(e) and assembling a sequence for each of the array of target polynucleotides from the stored identity and position of the imaging tags, optionally as a homogeneous or one pot reaction. Additional methods of sequencing target polynucleotides are described herein.

Abstract: Methods of detecting, probing, mapping and directed sequencing of target nucleic acids are provided using a guide RNA and a Cas9 protein. Methods for detecting the binding of the guide RNA/Cas9 complex to a target nucleic acid where the guide RNA includes a 3? tail sequence that can hybridize to a probe are provided. Methods for detecting the binding of the guide RNA/Cas9 complex to a target nucleic acid where the complex is physically detected are provided.

Abstract: The present invention relates to a method of sequencing a target polynucleotide by enzymatic and/or chemical means. The sequencing method includes a method for characterizing multiple alleles in a sample, a method of calculating confidence levels in ascertained sequences, a method for comparing polynucleotide sequences and a method of resolving ambiguities in a polynucleotide sequence. It also provides methods for appropriately preparing samples, for immobilising template molecules, for organising the template molecules and to conduct the sequencing of many molecules in parallel. The method involves analysing molecules as members of an array. Many target polynucleotides or many segments of a single target polynucleotide can be sequenced simultaneously. In a preferred embodiment the method involves analysing individual molecules within an array and base calls are based on the signals from two or more molecules. A method to prevent non-specific signal in sequencing is also provided.

Abstract: The present invention relates to a method of sequencing a target polynucleotide by enzymatic and/or chemical means. The sequencing method includes a method for characterizing multiple alleles in a sample, a method of calculating confidence levels in ascertained sequences, a method for comparing polynucleotide sequences and a method of resolving ambiguities in a polynucleotide sequence. It also provides methods for appropriately preparing samples, for immobilising template molecules, for organising the template molecules and to conduct the sequencing of many molecules in parallel. The method involves analysing molecules as members of an array. Many target polynucleotides or many segments of a single target polynucleotide can be sequenced simultaneously. In a preferred embodiment the method involves analysing individual molecules within an array and base calls are based on the signals from two or more molecules. A method to prevent non-specific signal in sequencing is also provided.