pET28a-T5-ASG_ClpXP
(Plasmid #153299)

• Purpose: Expresses acoustic sensor genes or ClpXP-sensitive gas vesicles in E. coli Nissle 1917
• Depositing Lab: Mikhail Shapiro
• Publication: Lakshmanan et al Nat Chem Biol. 2020 Sep;16(9):988-996. doi: 10.1038/s41589-020-0591-0. Epub 2020 Jul 13.

Backbone

• Vector backbone: pET28a
• Backbone manufacturer: Novagen
• Backbone size w/o insert (bp): 5189
• Total vector size (bp): 11983
• Vector type: Bacterial Expression, Synthetic Biology

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): NEB Turbo
• Growth instructions: Following transformation, rescue in SOC broth for 1 hour at 37C before transferring the rescued cells (entire volume) into overnight growth media (LB+ 50 ug/mL Kanamycin + 2% glucose) for growth at 37C. 2% glucose is optional, but recommend to prevent any leaky protein expression from the constitutive T5 promoter driving the ARG gene. Usually a 40% stock solution of glucose is prepared and diluted in the media to the final working concentration of 2%.
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: ASG
• Alt name: Acoustic Sensor Gene cluster
• Alt name: Hybrid gas vesicle gene cluster
• Species: Anabaena flos-aquae and Bacillus megaterium
• Insert Size (bp): 6794
• Promoter: T5

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: custom primers
• 3′ sequencing primer: T7 terminator reverse

Resource Information

• Supplemental Documents:
  • pET28a_T5-ASG_ClpXP.gb

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry