Open AccessOne-Enzyme Reverse Transcription qPCR Using Taq DNA Polymerase
Author(s) -
Sanchita Bhadra,
Andre C. Maranhao,
Inyup Paik,
Andrew D. Ellington
Publication year - 2020
Publication title -
biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.43
H-Index - 253
eISSN - 1520-4995
pISSN - 0006-2960
DOI - 10.1021/acs.biochem.0c00778
Subject(s) - taqman , taq polymerase , reverse transcriptase , dna polymerase , polymerase , microbiology and biotechnology , biology , real time polymerase chain reaction , dna , hot start pcr , virology , polymerase chain reaction , thermus aquaticus , genetics , nested polymerase chain reaction , gene
Taq DNA polymerase, one of the first thermostable DNA polymerases to be discovered, has been typecast as a DNA-dependent DNA polymerase commonly employed for PCR. However, Taq polymerase belongs to the same DNA polymerase superfamily as the Molony murine leukemia virus reverse transcriptase and has in the past been shown to possess reverse transcriptase activity. We report optimized buffer and salt compositions that promote the reverse transcriptase activity of Taq DNA polymerase and thereby allow it to be used as the sole enzyme in TaqMan RT-qPCRs. We demonstrate the utility of Taq-alone RT-qPCRs by executing CDC SARS-CoV-2 N1, N2, and N3 TaqMan RT-qPCR assays that could detect as few as 2 copies/μL of input viral genomic RNA.