pET28a-ValRS
(Plasmid #259970)

• Purpose: Expression vector for producing E. coli valyl-tRNA synthetase (ValRS), a component of the PURE cell-free transcription/translation system, in E. coli.
• Depositing Lab: Jonathan Calles
• Depositing Organization: b.next
  Located in the United States of America
• Publication: Protein Synthesis Using Recombinant Elements (PURE) cell-free transcription/translation system in E. coli (unpublished)

Backbone

• Vector backbone: pET28a
• Backbone manufacturer: Novagen / MilliporeSigma
• Backbone size w/o insert (bp): 5246
• Vector type: Bacterial Expression

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Growth instructions: For protein expression, transform into BL21(DE3) and induce with IPTG. DH5alpha is recommended for plasmid propagation/distribution.
• Copy number: Low Copy

Gene/Insert

• Gene/Insert name: ValRS
• Species: E. coli (Ec)
• Insert Size (bp): 2853
• Entrez Gene: valS (a.k.a. b4258, ECK4251, JW4215, val-act)
• Promoter: T7
• Tag / Fusion Protein:
  • His6 (C terminal on insert)

Cloning Information

• Cloning method: Gene Synthesis

Resource Information

• A portion of this plasmid was derived from a plasmid made by: Synthesis

Terms and Licenses

• Academic/Nonprofit Terms:
  • OpenMTA
• Industry Terms:
  • OpenMTA

Depositor Comments

Expression is lac-inducible (IPTG). This plasmid is an original b.next design for implementing the PURE cell-free system, first described by Shimizu et al. (Nat Biotechnol 2001, PMID 11479568). It can be produced using the OnePot method from Lavickova & Maerkl (ACS Synth Biol 2019, PMID 30632751).

Protocols for using these plasmids, along with validation data documented as Developer Notes, are linked in the Resource Information section.

For additional information about Nucleus, the open developer platform for synthetic cells, see: https://bnext.bio/nucleus.