SOLUTION-PHASE, TRANS-ACTIVATED REPORTER SYSTEMS FOR USE IN CRISPR-BASED NUCLEIC ACID SEQUENCE DETECTIONS
摘要:
Embodiments disclosed herein include devices, methods, and systems for direct, selective, and sensitive detection of single-stranded and double-stranded target nucleic acid sequences from various sources in a solution-based system. When activated by binding a target nucleic acid sequence, the Cas protein cleaves a tether separating a reporter molecule from a capture moiety. The capture moiety can then be used to remove, localize, or sequester uncleaved molecule containing intact tethers. In some embodiments, the systems, methods, and devices may include a filter, a membrane, or other molecules that may help to separate the tethered and untethered reporter molecules and/or capture the reporter molecules. These devices, systems, and techniques allow a user to rapidly process samples that may contain the target nucleic acid, in some cases, without needing to amplify the target sequences, and without the need for sophisticated or expensive laboratory equipment. These devices and methods may be used to assay a wide variety of samples and target nucleic acid sources, for the presence or absence of a specific target sequences. Compositions and kits, useful in practicing these methods, for example detecting a target RNA or DNA in a biological sample, are also described.
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