LOCUS       sequence-263054-        3951 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pFE carrying a human carbonic anhydrase II gene (Uniprot: P00918).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3951)
  AUTHORS   Desmarais JJ, Flamholz AI, Blikstad C, Dugan EJ, Laughlin TG, 
            Oltrogge LM, Chen AW, Wetmore K, Diamond S, Wang JY, Savage DF
  TITLE     DABs are inorganic carbon pumps found throughout prokaryotic phyla.
  JOURNAL   Nat Microbiol. 2019 Aug 12. pii: 10.1038/s41564-019-0520-8. doi: 
            10.1038/s41564-019-0520-8.
  PUBMED    31406332
REFERENCE   2  (bases 1 to 3951)
  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: "Nat 
            Microbiol. 2019 Aug 12. pii: 10.1038/s41564-019-0520-8. doi: 
            10.1038/s41564-019-0520-8."
COMMENT     Sequence Label: pFE-hCA
FEATURES             Location/Qualifiers
     source          1..3951
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     290..309
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     542..560
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(581..599,600..605,606..622,623..628,629..654)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    581..599
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    606..624
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             664..675
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     RBS             686..697
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1484..1501
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1556..1642
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1815..2438)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     2346..2365
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(2439..2543)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2719..3307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2799..2818)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(3395..3489)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3520..3951,1..363))
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3631..3650)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 ccgcattgca tcagccatga tggatacttt ctcggcagga gcaaggtgag atgacaggag
       61 atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag tgacaacgtc
      121 gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc
      181 ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg
      241 cgctgacagc cggaacacgg cggcatcaga gcagccgatt gtctgttgtg cccagtcata
      301 gccgaatagc ctctccaccc aagcggccgg agaacctgcg tgcaatccat cttgttcaat
      361 catgcgaaac gatcctcatc ctgtctcttg atcagatctt gatcccctgc gccatcagat
      421 ccttggcggc aagaaagcca tccagtttac tttgcagggc ttcccaacct taccagaggg
      481 cgccccagct ggcaattccg acgtctaaga aaccattatt atcatgacat taacctataa
      541 aaataggcgt atcacgaggc cctttcgtct tcacctcgag tccctatcag tgatagagat
      601 tgacatccct atcagtgata gagatactga gcacatcagc aggacgcact gaccgaattc
      661 attaaagagg agaaaggtac cgtccaaaga ggagaaaggt accatgtcgc atcactgggg
      721 ttatggaaaa cacaacggcc cggagcattg gcacaaagat tttcccattg cgaaggggga
      781 acgccaatcc cctgtcgaca ttgacaccca cacggcaaag tatgacccta gccttaaacc
      841 attatctgta agctacgatc aagcgacttc cctgcgtatc ttaaacaatg ggcatgcctt
      901 taatgtggag ttcgatgata gccaggacaa ggccgttctg aaagggggtc cacttgatgg
      961 cacctaccgc ctgatccaat tccactttca ctgggggagt ctggacggcc aaggctctga
     1021 gcatacagtt gacaagaaga agtacgccgc agaacttcac ctggttcatt ggaataccaa
     1081 gtatggagac ttcggcaaag ctgtgcaaca accagacggc ttggcggtgt tgggtatctt
     1141 ccttaaagtg ggatctgcca aacctgggtt acagaaggtg gtggatgtcc ttgatagtat
     1201 taagactaag ggaaaaagtg ctgactttac caactttgat ccgcgtggtt tattaccgga
     1261 gagcttagat tattggactt atccgggttc attaaccaca cctcccttac tggaatgcgt
     1321 gacttggatt gtcctgaaag agcccatttc cgtgtcaagc gagcaagtct taaagtttcg
     1381 caaacttaat ttcaatggtg aaggggaacc agaagaactt atggtggaca actggcgccc
     1441 ggcgcagccc ttaaaaaatc gccagatcaa agcgtcattc aaacaccatc accatcacca
     1501 ttgaagcgga attcctgcag cccgggggat cccatggtac gcgtgctaga ggcatcaaat
     1561 aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa
     1621 cgctctcctg agtaggacaa atccgccgcc gctagcagtt cgtagacgga aacaaacgca
     1681 gaatccaagc aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     1741 tttcaatatt attgaagcat ttatcagggt tatgtccact gggttcgtgc cttcatccgg
     1801 atatccagga actgttaaga cccactttca catttaagtt gtttttctaa tccgcatatg
     1861 atcaattcaa ggccgaataa gaaggctggc tctgcacctt ggtgatcaaa taattcgata
     1921 gcttgtcgta ataatggcgg catactatca gtagtaggtg tttccctttc ttctttagcg
     1981 acttgatgct cttgatcttc caatacgcaa cctaaagtaa aatgccccac agcgctgagt
     2041 gcatataatg cattctctag tgaaaaacct tgttggcata aaaaggctaa ttgattttcg
     2101 agagtttcat actgtttttc tgtaggccgt gtacctaaat gtacttttgc tccatcgcga
     2161 tgacttagta aagcacatct aaaactttta gcgttattac gtaaaaaatc ttgccagctt
     2221 tccccttcta aagggcaaaa gtgagtatgg tgcctatcta acatctcaat ggctaaggcg
     2281 tcgagcaaag cccgcttatt ttttacatgc caatacaatg taggctgctc tacacctagc
     2341 ttctgggcga gtttacgggt tgttaaacct tcgattccga cctcattaag cagctctaat
     2401 gcgctgttaa tcactttact tttatctaat ctagacatac tcttcctttt tcaatattat
     2461 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     2521 aataaacaaa taggggttcc gcgtttccct cgacaattcg cgctagacct agggcgttcg
     2581 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     2641 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     2701 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     2761 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     2821 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     2881 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     2941 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     3001 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     3061 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     3121 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     3181 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     3241 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     3301 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     3361 acgttaaggg attttggtca tgactagtgc ttggattctc accaataaaa aacgcccggc
     3421 ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca
     3481 acaggagtcc aagcgagctc tcgaacccca gagtcccgct cagaagaact cgtcaagaag
     3541 gcgatagaag gcgatgcgct gcgaatcggg agcggcgata ccgtaaagca cgaggaagcg
     3601 gtcagcccat tcgccgccaa gctcttcagc aatatcacgg gtagccaacg ctatgtcctg
     3661 atagcggtcc gccacaccca gccggccaca gtcgatgaat ccagaaaagc ggccattttc
     3721 caccatgata ttcggcaagc aggcatcgcc atgggtcacg acgagatcct cgccgtcggg
     3781 catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg agcccctgat gctcttcgtc
     3841 cagatcatcc tgatcgacaa gaccggcttc catccgagta cgtgctcgct cgatgcgatg
     3901 tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc gtatgcagcc g
//
