pInsert-PhoCl-RER
(Plasmid #219676)

• Purpose: Caging at cytosolic C-ter of POI: mScarletCD4-PhoCl-RER in pTREtight2
• Depositing Lab: Helge Ewers
• Publication: Kashyap et al Nat Methods. 2024 Mar 8. doi: 10.1038/s41592-024-02204-x.

Backbone

• Vector backbone: pTREtight2
• Vector type: Mammalian Expression

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: CD4mScarlet-PhoCl-RER
• Promoter: TRE
• Tag / Fusion Protein:
  • mScarlet and PhoCl

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: T7
• 3′ sequencing primer: T7 Term

Resource Information

• Supplemental Documents:
  • pInsert-PhoCl-RER.gb

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Industry MTA

Depositor Comments

PhoCl2c-RER can be cut out with HindIII and put in the desired vector with
the POI. can be done easily with gibson cloning in one step.

All the plasmids with pTRE vectors have the POI under tetracycline
promoter and hence when transfected needs to be transfected with another
plasmid, rtTA3(Sara knows) in the presence of doxycycline. or it can be
transfected in cells stably expressing the tetracycline-on transactivator
(rtTA3). rtTA3 is the transactivator that binds with doxycycline and sits
at the promoter for initiation of transcription.

While cloning please make sure you include signal peptide in the beginning
if needed. This depends on the specific POI.