LOCUS       sequence-323480-        4747 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Arabinose-inducible expression of green fluorescent protein encoding
            zero amber codons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4747)
  AUTHORS   Ho JML, Miller CA, Parks SE, Mattia JR, Bennett MR
  TITLE     A suppressor tRNA-mediated feedforward loop eliminates leaky gene 
            expression in bacteria.
  JOURNAL   Nucleic Acids Res. 2021 Mar 18;49(5):e25. doi: 10.1093/nar/gkaa1179.
  PUBMED    33290521
REFERENCE   2  (bases 1 to 4747)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res."; date: "2021-03-18"; volume: "49(5):e25. doi"; pages: " 
            10.1093/nar/gkaa1179"
COMMENT     Sequence Label: pJH25_pBad_AraC_Para-sfGFP.0TAG
FEATURES             Location/Qualifiers
     source          1..4747
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     208..227
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             join(323..325,326..1036)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             1043..1060
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1137..1154)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1307..1393
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1485..1512
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1531..1622
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1623..1691,1692..2483)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1841..1860)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2654..3242
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3143..3162
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3407..3424
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3439..3579
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3505..3524)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3665..3687
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3843..4721)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccta ggatgagcaa gggcgaagaa ctgtttacgg gcgtggtgcc
      361 gattctggtg gaactggatg gtgatgtcaa tggtcacaaa ttcagcgtgc gcggcgaagg
      421 tgaaggcgat gcaaccaatg gtaaactgac gctgaagttt atttgcacca cgggtaaact
      481 gccggttccg tggccgaccc tggtcaccac gctgacgtat ggtgttcagt gtttcagtcg
      541 ttacccggat cacatgaaac gccacgactt tttcaagtcc gcgatgccgg aaggttatgt
      601 ccaagaacgt accatctcat ttaaagatga cggcacctac aaaacgcgcg ccgaagtgaa
      661 attcgaaggt gatacgctgg ttaaccgtat tgaactgaaa ggcatcgatt ttaaggaaga
      721 cggtaatatt ctgggccata aactggaata taacttcaat tcgcacaacg tgtacatcac
      781 cgcagataag cagaagaacg gtatcaaggc taacttcaag atccgccata atgtggaaga
      841 tggcagcgtt caactggccg accactatca gcaaaacacc ccgattggtg atggcccggt
      901 cctgctgccg gacaatcatt acctgagcac gcagtctgtg ctgagtaaag atccgaacga
      961 aaagcgtgac cacatggtcc tgctggaatt cgtgaccgcg gccggcatca cgcacggtat
     1021 ggacgaactg tataaaggct cacatcatca tcatcatcat tgactcgaga tctgcagctg
     1081 gtaccatatg ggaattcgaa gcttggctgt tttggcggat gagagaagat tttcagcctg
     1141 atacagatta aatcagaacg cagaagcggt ctgataaaac agaatttgcc tggcggcagt
     1201 agcgcggtgg tcccacctga ccccatgccg aactcagaag tgaaacgccg tagcgccgat
     1261 ggtagtgtgg ggtctcccca tgcgagagta gggaactgcc aggcatcaaa taaaacgaaa
     1321 ggctcagtcg aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcct
     1381 gagtaggaca aatccgccgg gagcggattt gaacgttgcg aagcaacggc ccggagggtg
     1441 gcgggcagga cgcccgccat aaactgccag gcatcaaatt aagcagaagg ccatcctgac
     1501 ggatggcctt tttgcgtttc tacaaactct tttgtttatt tttctaaata cattcaaata
     1561 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     1621 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     1681 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     1741 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     1801 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     1861 cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     1921 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     1981 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     2041 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     2101 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     2161 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     2221 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     2281 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     2341 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     2401 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     2461 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     2521 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2581 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     2641 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     2701 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     2761 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     2821 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     2881 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     2941 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3001 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3061 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3121 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3181 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3241 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctggcc
     3301 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     3361 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     3421 cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     3481 acaccgcata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
     3541 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
     3601 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     3661 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     3721 cagatcaatt cgcgcgcgaa ggcgaagcgg catgcataat gtgcctgtca aatggacgaa
     3781 gcagggattc tgcaaaccct atgctactcc gtcaagccgt caattgtctg attcgttacc
     3841 aattatgaca acttgacggc tacatcattc actttttctt cacaaccggc acggaactcg
     3901 ctcgggctgg ccccggtgca ttttttaaat acccgcgaga aatagagttg atcgtcaaaa
     3961 ccaacattgc gaccgacggt ggcgataggc atccgggtgg tgctcaaaag cagcttcgcc
     4021 tggctgatac gttggtcctc gcgccagctt aagacgctaa tccctaactg ctggcggaaa
     4081 agatgtgaca gacgcgacgg cgacaagcaa acatgctgtg cgacgctggc gatatcaaaa
     4141 ttgctgtctg ccaggtgatc gctgatgtac tgacaagcct cgcgtacccg attatccatc
     4201 ggtggatgga gcgactcgtt aatcgcttcc atgcgccgca gtaacaattg ctcaagcaga
     4261 tttatcgcca gcagctccga atagcgccct tccccttgcc cggcgttaat gatttgccca
     4321 aacaggtcgc tgaaatgcgg ctggtgcgct tcatccgggc gaaagaaccc cgtattggca
     4381 aatattgacg gccagttaag ccattcatgc cagtaggcgc gcggacgaaa gtaaacccac
     4441 tggtgatacc attcgcgagc ctccggatga cgaccgtagt gatgaatctc tcctggcggg
     4501 aacagcaaaa tatcacccgg tcggcaaaca aattctcgtc cctgattttt caccaccccc
     4561 tgaccgcgaa tggtgagatt gagaatataa cctttcattc ccagcggtcg gtcgataaaa
     4621 aaatcgagat aaccgttggc ctcaatcggc gttaaacccg ccaccagatg ggcattaaac
     4681 gagtatcccg gcagcagggg atcattttgc gcttcagcca tacttttcat actcccgcca
     4741 ttcagag
//
