pAS_4xMma PylT_FLAG-Mma PylRS AF
(Plasmid #140023)

• Purpose: Plasmid with 4xMmaPylT cassette and MmaPylRS AF for amber suppression; for transient or stable piggyBac-mediated integration
• Depositing Lab: Simon Elsaesser
• Publication: Meineke et al ACS Chem Biol. 2018 Sep 27. doi: 10.1021/acschembio.8b00571.

Backbone

• Vector backbone: pAS
• Vector type: Mammalian Expression
• Selectable markers: Puromycin

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: Mma PylRS
• Alt name: 4xh7SK-Mma PylT cassette
• Species: Methanosarcina mazei
• Insert Size (bp): 2619
• Mutation: Y271A, Y349F
• Entrez Gene: pylS (a.k.a. MSMAC_RS17105, MSMAC_3241)
• Promoter: EF1
• Tag / Fusion Protein:
  • FLAG (N terminal on insert)

Cloning Information

• Cloning method: Ligation Independent Cloning

Resource Information

• Supplemental Documents:
  • pas_4xmmapylt_mmapylrs-af.gb
• Articles Citing this Plasmid:
  • 2 References

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • piggyBac Limited Use Label License
• Industry Terms:
  • Not Available to Industry

Depositor Comments

5' Cloning site: NheI
3' Cloning site: SalI

The AF mutant (originally Y306A, Y384F in MbPylRS) has been described in:

Yanagisawa, T., Ishii, R., Fukunaga, R., Kobayashi, T., Sakamoto, K. and Yokoyama, S. 2008. Multistep engineering of pyrrolysyl-tRNA synthetase to genetically encode N(epsilon)-(o-azidobenzyloxycarbonyl) lysine for site-specific protein modification. Chemistry & Biology. PubMed: https://www.ncbi.nlm.nih.gov/pubmed/19022179