PNA-Assisted Rolling Circle Amplification for Detection of DNA Marker Sequences in Human Cells
2016
Presented here are the methods used for detection of short, single-copy DNA sequences within human cells. These methods rely on (1) peptide nucleic acid (PNA) invasion of the target double-stranded DNA site to locally open a small single-stranded region within long DNA duplex, (2) site-specific formation of padlock probe, and (3) its rolling circle amplification for signal magnification. The signal detection is accomplished via either fluorescent in situ hybridization (FISH), flow cytometry (fluorescence-activated cell sorting, FACS), or droplet-based microfluidics. This methodology allows for sensitive and specific detection of DNA marker sequences without the need for DNA extraction and purification, making it possible to identify small genomic variations within individual human cells.
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