Nanoliter scale PCR with TaqMan detection | Zendy

Olga Kalinina | Zendy

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Nanoliter scale PCR with TaqMan detection

Author(s) -

Olga Kalinina

Publication year - 1997

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/25.10.1999

Subject(s) - taqman , biology , microbiology and biotechnology , serial dilution , fluorescence , dna , genomic dna , polymerase chain reaction , taq polymerase , replicate , real time polymerase chain reaction , chromatography , computational biology , genetics , polymerase , gene , chemistry , medicine , thermus aquaticus , statistics , alternative medicine , physics , mathematics , pathology , quantum mechanics

We monitored PCR in volumes of the order of 10 nl in glass microcapillaries using a fluorescence energy transfer assay in which fluorescence increases if product is made due to template-dependent nucleolytic degradation of an internally quenched probe (TaqMan assay). This assay detected single starting template molecules in dilutions of genomic DNA. The results suggest that it may be feasible to determine the number of template molecules in a sample by counting the number of positive PCRs in a set of replicate reactions using terminally diluted sample. Since the assay system is closed and potentially automatable, it has promise for clinical applications.

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