Sequencing method
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United States of America Patent
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Sep 26, 2017
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app pub date -
Oct 4, 2013
filing date -
Oct 4, 2012
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Abstract
Disclosed is a method for sequencing a polynucleotide analyte comprising: •a. generating a stream of droplets containing a single nucleotide wherein the order of single nucleotides in the droplet stream corresponds to the sequence of nucleotides in the analyte; •b. introducing into each droplet a plurality of biological probe types each type comprising a different label in an undetectable state and being adapted to capture a different single nucleotide; •c. causing the single nucleotide contained in the droplet to bind to its complementary probe and •d. causing the label to be released from the probe that has bound the nucleotide in a detectable state. The probe is a dumbbell shaped probe comprising fluorescent donor and quencher labels and a single nucleotide gap. After gap repair by a polymerase and a ligase, a restriction enzyme recognition site is cleaved by a restriction enzyme, followed by exonuclease digestion to release the labels.
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