pF420
(Plasmid #123659)

• Purpose: Produces cofactor F420 in E. coli
• Depositing Lab: Colin Jackson
• Publication: Bashiri et al Nat Commun. 2019 Apr 5;10(1):1558. doi: 10.1038/s41467-019-09534-x.

Backbone

• Vector backbone: pSB1C3
• Backbone manufacturer: iGEM
• Backbone size w/o insert (bp): 2070
• Total vector size (bp): 8808
• Modifications to backbone: Additionally expresses TetR cassette BBa_K145201.
• Vector type: Bacterial Expression, Synthetic Biology

Growth in Bacteria

• Bacterial Resistance(s): Chloramphenicol, 25 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): Top10
• Copy number: High Copy

Gene/Insert 1

• Gene/Insert name: CofD
• Alt name: FbiA
• Alt name: EPPG:Fo phosphoenolpyruvyl transferase
• Species: Methanosarcina mazei
• Insert Size (bp): 909
• GenBank ID: MM_RS09715
• Promoter: BBa_R0040

Cloning Information for Gene/Insert 1

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (not destroyed)
• 3′ cloning site: SpeI (destroyed during cloning)
• 5′ sequencing primer: GTTCACCACGTTTACCAATC
• 3′ sequencing primer: AACCAGCACATCACCGTCAC

Gene/Insert 2

• Gene/Insert name: FbiB
• Alt name: CofB
• Alt name: F420-0:γ-glutamate ligase
• Species: Mycobacterium smegmatis
• Insert Size (bp): 1362
• GenBank ID: MSMEG_1892
• Promoter: BBa_R0040

Cloning Information for Gene/Insert 2

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (destroyed during cloning)
• 3′ cloning site: SpeI (destroyed during cloning)
• 5′ sequencing primer: AACCAGCACATCACCGTCAC
• 3′ sequencing primer: ACCTGTTTTGGCTGGGCACC

Gene/Insert 3

• Gene/Insert name: FbiD
• Alt name: CofC
• Alt name: phosphoenolpyruvate guanylyltransferase
• Species: Mycobacterium smegmatis
• Insert Size (bp): 663
• GenBank ID: MSMEG_2392
• Promoter: BBa_R0040

Cloning Information for Gene/Insert 3

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (destroyed during cloning)
• 3′ cloning site: SpeI (destroyed during cloning)
• 5′ sequencing primer: TGCAGCAAGTGATGGTAGTG
• 3′ sequencing primer: None

Gene/Insert 4

• Gene/Insert name: FbiC
• Alt name: CofGH
• Alt name: FO Synthase
• Species: Mycobacterium smegmatis
• Insert Size (bp): 2622
• GenBank ID: MSMEG_5126
• Promoter: BBa_R0040

Cloning Information for Gene/Insert 4

• Cloning method: Restriction Enzyme
• 5′ cloning site: EcoRI (destroyed during cloning)
• 3′ cloning site: SpeI (not destroyed)
• 5′ sequencing primer: TAAGGAGCCCCCATAATGCG
• 3′ sequencing primer: ATGGTCATGTTGATCAGCCT

Resource Information

• Supplemental Documents:
  • pF420.gb

Terms and Licenses

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

Depositor Comments

Full set of sequencing primers:

U4965CK070-6-SEQ1R AACCAGCACATCACCGTCAC
U4965CK070-6-SEQ2R CGGTCAGGGCTTCACCAACT
U4965CK070-6-SEQ3 ACCTGTTTTGGCTGGGCACC
U4965CK070-6-SEQ4 TGCAGCAAGTGATGGTAGTG
U4965CK070-6-SEQ5 TAAGGAGCCCCCATAATGCG
U4965CK070-6-SEQ6N GAGTTGGAGCGAACTGAGTC
U4965CK070-6-SEQ7 ACCGCCTAATCTGGTTAGCC
U4965CK070-6-SEQ8 CCTGGTGGTTGTAGTGATGA
U4965CK070-6-SEQ9 ATGGTCATGTTGATCAGCCT
U4965CK070-6-SEQ0.5r GTTCACCACGTTTACCAATC