Open AccessAgarose gel electrophoresis to assess PCR product yield: comparison with spectrophotometry, fluorometry and qPCR
Author(s) -
Patrick Wittmeier,
Susanne Hummel
Publication year - 2022
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/btn-2021-0094
Subject(s) - agarose , agarose gel electrophoresis , amplicon , chromatography , fluorescence spectroscopy , gel electrophoresis , fluorescence , electrophoresis , spectrophotometry , chemistry , microbiology and biotechnology , polymerase chain reaction , biology , dna , biochemistry , gene , physics , quantum mechanics
Agarose gel electrophoresis is a relatively easy to use method, commonly applied to evaluate PCR reaction success. Intercalating agents or dyes are used to visualize the amplified fragments. However, it is uncertain to what extent the brightness of bands is informative about the concentration of the amplicons. To more closely examine the suitability of agarose gel electrophoresis to assess PCR product yield, we quantified the brightness of bands on a gel and compared these data with the results from spectrophotometry, fluorometry and qPCR. Evaluation of the results suggests that assessment of the relative quantity of amplicons by band brightness is precise enough even for post-PCR analysis steps requiring PCR product concentrations within a certain range to function properly.