Efficient tRNA degradation and quantification in Escherichia coli cell extract using RNase-coated magnetic beads: A key step toward codon emancipation.
Biotechnol Prog
; 33(5): 1401-1407, 2017 Sep.
Article
en En
| MEDLINE
| ID: mdl-28593644
Emancipating sense codons toward a minimized genetic code is of significant interest to science and engineering. A key approach toward sense codon emancipation is the targeted in vitro removal of native tRNA. However, challenges remain such as the insufficient depletion of tRNA in lysate-based in vitro systems and the high cost of the purified components system (PURE). Here we used RNase-coated superparamagnetic beads to efficiently degrade E. coli endogenous tRNA. The presented method removes >99% of tRNA in cell lysates, while partially preserving cell-free protein synthesis activity. The resulting tRNA-depleted lysate is compatible with in vitro-transcribed synthetic tRNA for the production of peptides and proteins. Additionally, we directly measured residual tRNA using quantitative real-time PCR. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1401-1407, 2017.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Ribonucleasa Pancreática
/
ARN de Transferencia
/
Extractos Celulares
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Escherichia coli
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Biología Sintética
Límite:
Animals
Idioma:
En
Revista:
Biotechnol Prog
Asunto de la revista:
BIOTECNOLOGIA
Año:
2017
Tipo del documento:
Article
Pais de publicación:
Estados Unidos