Optimizing protein V untranslated region sequence in M13 phage for increased production of single-stranded DNA for origami.
Nucleic Acids Res
; 49(11): 6596-6603, 2021 06 21.
Article
em En
| MEDLINE
| ID: mdl-34110422
ABSTRACT
DNA origami requires long scaffold DNA to be aligned with the guidance of short staple DNA strands. Scaffold DNA is produced in Escherichia coli as a form of the M13 bacteriophage by rolling circle amplification (RCA). This study shows that RCA can be reconfigured by reducing phage protein V (pV) expression, improving the production throughput of scaffold DNA by at least 5.66-fold. The change in pV expression was executed by modifying the untranslated region sequence and monitored using a reporter green fluorescence protein fused to pV. In a separate experiment, pV expression was controlled by an inducer. In both experiments, reduced pV expression was correlated with improved M13 bacteriophage production. High-cell-density cultivation was attempted for mass scaffold DNA production, and the produced scaffold DNA was successfully folded into a barrel shape without compromising structural quality. This result suggested that scaffold DNA production throughput can be significantly improved by reprogramming the RCA in E. coli.
Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
Proteínas Virais
/
DNA de Cadeia Simples
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Bacteriófago M13
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Proteínas de Ligação a DNA
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article