The present invention relates to the detection of a target nucleic acid sequence by a PCE-NH (PTO Cleavage and Extension-Dependent Non-Hybridization) assay. The present invention adopts the occurrence of the inhibition of the hybridization between the HO with the CTO by the formation of the target-dependent extended duplex. Therefore, the present invention may detect target sequences even when the HO is not cleaved. In this regard, the design of the 5′-tagging portion of PTO, CTO and HO sequences may be readily performed and the conditions for reactions may be also easily established. In addition, the detection of the hybrid between the CTO and the HO may be performed in a different vessel from that for the extension of the CTO.

The present invention relates to the detection of a target nucleic acid sequence by a PCE-NH (PTO Cleavage and Extension-Dependent Non-Hybridization) assay. The present invention adopts the occurrence of the inhibition of the hybridization between the HO with the CTO by the formation of the target-dependent extended duplex. Therefore, the present invention may detect target sequences even when the HO is not cleaved. In this regard, the design of the 5′-tagging portion of PTO, CTO and HO sequences may be readily performed and the conditions for reactions may be also easily established. In addition, the detection of the hybrid between the CTO and the HO may be performed in a different vessel from that for the extension of the CTO.

This invention provides methods for characterizing the amounts of nucleic acids, including plus/minus determinations, the use of different constructs, the use of a library and a reference library. Expression may also be compared in two or more samples using the methods of this invention. Also provided are heterophasic arrays comprising labeled positive copies of nucleic acids hybridized to the array and labeled negative copies of nucleic acids hybridized to the array, in which the labeled positive copies are separately quantifiable from the labeled negative copies.

This invention provides methods for characterizing the amounts of nucleic acids, including plus/minus determinations, the use of different constructs, the use of a library and a reference library. Expression may also be compared in two or more samples using the methods of this invention. Also provided are heterophasic arrays comprising labeled positive copies of nucleic acids hybridized to the array and labeled negative copies of nucleic acids hybridized to the array, in which the labeled positive copies are separately quantifiable from the labeled negative copies.

The present invention provides an apparatus and a method for detecting the presence of and/or determining the amount of a label-free microRNA using an atomic force microscope. The method is extremely selective and/or ultrasensitive. In particular, the present invention provides a cantilever comprising a probe that selectively binds to a double strand of DNA/RNA hybrid complex. The probe comprises a hybrid binding domain (HBD) or a variant thereof.

The present invention provides an apparatus and a method for detecting the presence of and/or determining the amount of a label-free microRNA using an atomic force microscope. The method is extremely selective and/or ultrasensitive. In particular, the present invention provides a cantilever comprising a probe that selectively binds to a double strand of DNA/RNA hybrid complex. The probe comprises a hybrid binding domain (HBD) or a variant thereof.

The present invention provides a novel miRNA extraction method and a method for analyzing miRNA extracted by using said miRNA extraction method. According to the present invention, provided is, for example, a method for extracting miRNA from extracellular vesicles in a sample solution, by using a device capable of capturing extracellular vesicles, the miRNA extraction method comprising: an extracellular vesicle capturing step for capturing extracellular vesicles in a sample solution onto a device by bringing the sample solution and the device in contact with each other; and a miRNA extraction step for homogenizing the extracellular vesicles by bringing the device having captured the extracellular vesicles in contact with a homogenization liquid for extracellular vesicles to extract miRNA from the extracellular vesicle into the homogenization liquid.

The present invention provides a novel miRNA extraction method and a method for analyzing miRNA extracted by using said miRNA extraction method. According to the present invention, provided is, for example, a method for extracting miRNA from extracellular vesicles in a sample solution, by using a device capable of capturing extracellular vesicles, the miRNA extraction method comprising: an extracellular vesicle capturing step for capturing extracellular vesicles in a sample solution onto a device by bringing the sample solution and the device in contact with each other; and a miRNA extraction step for homogenizing the extracellular vesicles by bringing the device having captured the extracellular vesicles in contact with a homogenization liquid for extracellular vesicles to extract miRNA from the extracellular vesicle into the homogenization liquid.

The present invention is directed to a method of designing a plurality of capture oligonucleotide probes for use on a support to which complementary oligonucleotide probes will hybridize with little mismatch, where the plural capture oligonucleotide probes have melting temperatures within a narrow range. The present invention further relates to an oligonucleotide array comprising of a support with the plurality of oligonucleotide probes immobilized on the support, a method of using the support to detect single-base changes, insertions, deletions, or translocations in a plurality of target nucleotide sequences, and a kit for such detection, which includes the support on which the oligonucleotides have been immobilized.

The present invention is directed to a method of designing a plurality of capture oligonucleotide probes for use on a support to which complementary oligonucleotide probes will hybridize with little mismatch, where the plural capture oligonucleotide probes have melting temperatures within a narrow range. The present invention further relates to an oligonucleotide array comprising of a support with the plurality of oligonucleotide probes immobilized on the support, a method of using the support to detect single-base changes, insertions, deletions, or translocations in a plurality of target nucleotide sequences, and a kit for such detection, which includes the support on which the oligonucleotides have been immobilized.