Using Digital Polymerase Chain Reaction to Detect Single-Nucleotide Substitutions Induced by Genome Editing | Zendy

Yuichiro Miyaoka | Zendy; Amanda H. Chan | Zendy; Bruce R. Conklin | Zendy

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Using Digital Polymerase Chain Reaction to Detect Single-Nucleotide Substitutions Induced by Genome Editing

Author(s) -

Yuichiro Miyaoka,

Amanda H. Chan,

Bruce R. Conklin

Publication year - 2016

Publication title -

cold spring harbor protocols

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 0.674

H-Index - 51

eISSN - 1940-3402

pISSN - 1559-6095

DOI - 10.1101/pdb.prot086801

Subject(s) - digital polymerase chain reaction , genome editing , polymerase chain reaction , computational biology , genome , nucleotide , single nucleotide polymorphism , biology , genetics , chemistry , gene , genotype

This protocol is designed to detect single-nucleotide substitutions generated by genome editing in a highly sensitive and quantitative manner. It uses a combination of allele-specific hydrolysis probes and a new digital polymerase chain reaction (dPCR) technology called droplet digital PCR (ddPCR). ddPCR partitions a reaction into more than 10,000 nanoliter-scale water-in-oil droplets. As a result, each droplet contains only a few copies of the genome so that ddPCR is able to detect rare genome-editing events without missing them.

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