The present invention belongs to the fields of biomedical technology and molecular diagnosis. Disclosed is a high-throughput detection method for a rare mutation of a gene, comprising: designing specific probes; connecting Y-shaped universal linkers to a test DNA subjected to fragmentation processing, and performing amplification and enrichment of a target site by universal sequence combination of the specific probes and the linkers; performing genomic sequence alignment on sequences to be sequenced; sorting and analyzing said sequences at the same starting and ending positions, and filtering sequencing errors; and after the data filtering, the sequencing depth count of a reference allele of the target site being a, and the sequencing depth count of other alleles being b, and thus the actual mutation ratio of the site being b/(a+b). This technique can perform, by DNA fragmentation, universal linker connection, multiplex PCR amplification of specific primers and linker sequence primers, and high-throughput high-depth sequencing, enrichment and parallel sequencing on a plurality of sites to be tested.

The present invention belongs to the fields of biomedical technology and molecular diagnosis. Disclosed is a high-throughput detection method for a rare mutation of a gene, comprising: designing specific probes; connecting Y-shaped universal linkers to a test DNA subjected to fragmentation processing, and performing amplification and enrichment of a target site by universal sequence combination of the specific probes and the linkers; performing genomic sequence alignment on sequences to be sequenced; sorting and analyzing said sequences at the same starting and ending positions, and filtering sequencing errors; and after the data filtering, the sequencing depth count of a reference allele of the target site being a, and the sequencing depth count of other alleles being b, and thus the actual mutation ratio of the site being b/(a+b). This technique can perform, by DNA fragmentation, universal linker connection, multiplex PCR amplification of specific primers and linker sequence primers, and high-throughput high-depth sequencing, enrichment and parallel sequencing on a plurality of sites to be tested.

The invention relates to methods of multiplex detection of a plurality of target nucleic acids by contacting a sample with an acid reagent to remove bound nucleic acid detection systems, thereby allowing the same detection systems to be used again to detect different target nucleic acids and to provide for higher levels of multiplexing. The invention also relates to kits containing an acid reagent and optionally probes for detection of target nucleic acids.

The invention relates to methods of multiplex detection of a plurality of target nucleic acids by contacting a sample with an acid reagent to remove bound nucleic acid detection systems, thereby allowing the same detection systems to be used again to detect different target nucleic acids and to provide for higher levels of multiplexing. The invention also relates to kits containing an acid reagent and optionally probes for detection of target nucleic acids.

The present invention relates to a method and a DNA nanostructure for detecting a target structure. In particular, the present invention relates to a DNA nanostructure, which ensures a preferably linear dependence on the number of marker molecules and the measurement signal regardless of the physical arrangement of a plurality of such DNA nanostructures by virtue of the skilled selection of the shape of the DNA nanostructure and the placement of the marker molecules attached to it. The invention additionally relates to the use of said DNA nanostructures and other nanoreporters, preferably in combination with adapters which bind specifically to target molecules, in a method for quantifying a plurality of target molecules, preferably in a simultaneous manner, using a multiplex method.

The present invention relates to a method and a DNA nanostructure for detecting a target structure. In particular, the present invention relates to a DNA nanostructure, which ensures a preferably linear dependence on the number of marker molecules and the measurement signal regardless of the physical arrangement of a plurality of such DNA nanostructures by virtue of the skilled selection of the shape of the DNA nanostructure and the placement of the marker molecules attached to it. The invention additionally relates to the use of said DNA nanostructures and other nanoreporters, preferably in combination with adapters which bind specifically to target molecules, in a method for quantifying a plurality of target molecules, preferably in a simultaneous manner, using a multiplex method.

Provided herein are high-throughput, high-quality methods of consecutive in situ hybridization for analysis of the genome and/or transcriptome in an individual cell with single-molecule sensitivity. In particular, provided herein are methods comprising visualizing individual genomic loci or transcripts as single detectable signals (e.g., fluorescent spots) which remain in place during consecutive hybridization. In each cycle of consecutive hybridization, detectably labeled probes hybridize to the probe used in the previous cycle, and also introduce the binding sites for the probe of the following cycle. Through consecutive cycles of probe hybridization, imaging, and signal removal, different genomic loci or RNA species can be identified by unique detectable signal profiles (e.g., fluorescent spots with unique color sequences). The number of varied color sequences increases exponentially with the number of hybridization cycles, which enables the genome or transcriptome-wide analysis.

Provided herein are high-throughput, high-quality methods of consecutive in situ hybridization for analysis of the genome and/or transcriptome in an individual cell with single-molecule sensitivity. In particular, provided herein are methods comprising visualizing individual genomic loci or transcripts as single detectable signals (e.g., fluorescent spots) which remain in place during consecutive hybridization. In each cycle of consecutive hybridization, detectably labeled probes hybridize to the probe used in the previous cycle, and also introduce the binding sites for the probe of the following cycle. Through consecutive cycles of probe hybridization, imaging, and signal removal, different genomic loci or RNA species can be identified by unique detectable signal profiles (e.g., fluorescent spots with unique color sequences). The number of varied color sequences increases exponentially with the number of hybridization cycles, which enables the genome or transcriptome-wide analysis.

This present disclosure describes hybridization probes modularly constructed from several oligonucleotides with a pattern of designed complementary interactions, allowing the probes to sequence-specifically capture or analyze nucleic acid target sequences that are long and/or complex.

This present disclosure describes hybridization probes modularly constructed from several oligonucleotides with a pattern of designed complementary interactions, allowing the probes to sequence-specifically capture or analyze nucleic acid target sequences that are long and/or complex.