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Microscale Adaptation of In Vitro Transcription/Translation for High‐Throughput Screening of Natural Product Extract Libraries
Author(s) -
Lowell Andrew N.,
Santoro Nicholas,
Swaney Steven M.,
McQuade Thomas J.,
Schultz Pamela J.,
Larsen Martha J.,
Sherman David H.
Publication year - 2015
Publication title -
chemical biology and drug design
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.59
H-Index - 77
eISSN - 1747-0285
pISSN - 1747-0277
DOI - 10.1111/cbdd.12614
Subject(s) - natural product , high throughput screening , in vitro , computational biology , biology , translation (biology) , small molecule , chemistry , biochemistry , messenger rna , gene
Novel antimicrobials that effectively inhibit bacterial growth are essential to fight the growing threat of antibiotic resistance. A promising target is the bacterial ribosome, a 2.5 MDa organelle susceptible to several biorthogonal modes of action used by different classes of antibiotics. To promote the discovery of unique inhibitors, we have miniaturized a coupled transcription/translation assay using E. coli and applied it to screen a natural product library of ~30 000 extracts. We significantly reduced the scale of the assay to 2 μ L in a 1536‐well plate format and decreased the effective concentration of costly reagents. The improved assay returned 1327 hits (4.6% hit rate) with % CV and Z′ values of 8.5% and 0.74, respectively. This assay represents a significant advance in molecular screening, both in miniaturization and its application to a natural product extract library, and we intend to apply it to a broad array of pathogenic microbes in the search for novel anti‐infective agents.