The invention relates to control compositions for sequencing and for chemical analyses, such as analytical chemistry analyses. More particularly, the invention relates to control compositions for sequencing and for chemical analyses having at least one barcode sequence fragment and at least one universal sequence fragment, and to methods of their use.

The invention relates to control compositions for sequencing and for chemical analyses, such as analytical chemistry analyses. More particularly, the invention relates to control compositions for sequencing and for chemical analyses having at least one barcode sequence fragment and at least one universal sequence fragment, and to methods of their use.

The present disclosure relates in some aspects to methods for analyzing a target nucleic acid in a biological sample. In some aspects, provided herein are methods and compositions for detecting a region of interest in a target nucleic acid, wherein hybridization between an interrogatory region of a probe and a region of interest of the target nucleic acid is blocked by a blocking strand unless the interrogatory region is complementary to the region of interest. In some aspects, the methods provided herein increase specificity of detecting a region of interest in a target nucleic acid (e.g., a SNP in an RNA molecule). In some aspects, the presence, amount, and/or identity of a region of interest in a target nucleic acid is analyzed in situ. Also provided are polynucleotides, sets of polynucleotides, compositions, and kits for use in accordance with the methods, for example for RNA-targeting padlock probe-mediated SNP detection.

The present disclosure relates in some aspects to methods for analyzing a target nucleic acid in a biological sample. In some aspects, provided herein are methods and compositions for detecting a region of interest in a target nucleic acid, wherein hybridization between an interrogatory region of a probe and a region of interest of the target nucleic acid is blocked by a blocking strand unless the interrogatory region is complementary to the region of interest. In some aspects, the methods provided herein increase specificity of detecting a region of interest in a target nucleic acid (e.g., a SNP in an RNA molecule). In some aspects, the presence, amount, and/or identity of a region of interest in a target nucleic acid is analyzed in situ. Also provided are polynucleotides, sets of polynucleotides, compositions, and kits for use in accordance with the methods, for example for RNA-targeting padlock probe-mediated SNP detection.

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.sub.F, both of the corresponding pair of earlier hybridisation steps H.sub.E comprise 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 H.sub.E comprise 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.

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.sub.F, both of the corresponding pair of earlier hybridisation steps H.sub.E comprise 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 H.sub.E comprise 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.

Embodiments of a method and/or system can include generating a set of target-associated molecules (e.g., spike-in molecules) associated with one or more biological targets; generating one or more spike-in mixtures based on processing the set of target-associated molecules with one or more samples including the one or more biological targets; performing one or more Sanger sequencing operations on the one or more spike-in mixtures; determining one or more abundance metrics based on chromatogram-related outputs from the one or more Sanger sequencing operations; and/or facilitating characterization of one or more medical conditions based on the one or more abundance metrics.

Embodiments of a method and/or system can include generating a set of target-associated molecules (e.g., spike-in molecules) associated with one or more biological targets; generating one or more spike-in mixtures based on processing the set of target-associated molecules with one or more samples including the one or more biological targets; performing one or more Sanger sequencing operations on the one or more spike-in mixtures; determining one or more abundance metrics based on chromatogram-related outputs from the one or more Sanger sequencing operations; and/or facilitating characterization of one or more medical conditions based on the one or more abundance metrics.

Digital assay system, including methods, apparatus, and compositions, for performing target assays in partitions each containing a generic reporter and a specific reporter for target amplification.

Digital assay system, including methods, apparatus, and compositions, for performing target assays in partitions each containing a generic reporter and a specific reporter for target amplification.