LOCUS       sequence-262995-        7317 bp    DNA     circular SYN 15-APR-2025
DEFINITION  pFE carrying a dabA2 gene (Uniprot: D0KWS7) with a D353A mutation 
            and a dabB2 gene (Uniprot: D0KWS8).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7317)
  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 7317)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-dabAB2 D353A
FEATURES             Location/Qualifiers
     source          1..7317
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     192..211
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        467..571
                     /gene="bla"
                     /label=AmpR promoter
     CDS             572..1195
                     /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     complement(645..664)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      complement(1368..1454)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     RBS             5701..5712
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(join(5722..5747,5748..5753,5754..5770,5771..5776,
                     5777..5795))
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    5752..5770
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5777..5795
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     complement(5816..5834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             6013..6807
                     /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(6067..6086)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6677..6696
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     terminator      6838..6932
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      join(7020..7317,1..291)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
       61 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
      121 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
      181 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
      241 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
      301 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
      361 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
      421 cgccgcagcc gaacgcccta ggtctagcgc gaattgtcga gggaaacgcg gaacccctat
      481 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
      541 aatgcttcaa taatattgaa aaaggaagag tatgtctaga ttagataaaa gtaaagtgat
      601 taacagcgca ttagagctgc ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact
      661 cgcccagaag ctaggtgtag agcagcctac attgtattgg catgtaaaaa ataagcgggc
      721 tttgctcgac gccttagcca ttgagatgtt agataggcac catactcact tttgcccttt
      781 agaaggggaa agctggcaag attttttacg taataacgct aaaagtttta gatgtgcttt
      841 actaagtcat cgcgatggag caaaagtaca tttaggtaca cggcctacag aaaaacagta
      901 tgaaactctc gaaaatcaat tagccttttt atgccaacaa ggtttttcac tagagaatgc
      961 attatatgca ctcagcgctg tggggcattt tactttaggt tgcgtattgg aagatcaaga
     1021 gcatcaagtc gctaaagaag aaagggaaac acctactact gatagtatgc cgccattatt
     1081 acgacaagct atcgaattat ttgatcacca aggtgcagag ccagccttct tattcggcct
     1141 tgaattgatc atatgcggat tagaaaaaca acttaaatgt gaaagtgggt cttaacagtt
     1201 cctggatatc cggatgaagg cacgaaccca gtggacataa ccctgataaa tgcttcaata
     1261 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttg
     1321 cttggattct gcgtttgttt ccgtctacga actgctagcg gcggcggatt tgtcctactc
     1381 aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc
     1441 tttcgtttta tttgatgcct ctagcacgcg taccatggga tcccccgggc tgcaggaatt
     1501 ccgcttcaga tcggttccgg tcgccagtca ccgtgggcat gacggcgatg cgggatgccg
     1561 ttgtcatcga tatgcaggat gtgcatccag cggttgttga ttaacgcgtt cagcatgtcg
     1621 tgaccggcaa taattttatc gatttctgca atgggcgcct caatgtacgc cgagagccgc
     1681 agcggttcgt gccgccactg ttcaccatcg tgcagcgatt gcatggcaag accaactcgc
     1741 aggtcgccgc cattgccttc aatcaccccg acggtgccgc cgaccacatt atgcagcact
     1801 ttattgcccg ccccttgatg cagattatcg accgtggagc cgtagtattg caggttgatc
     1861 cagtttgcga cgatcaaggg cgcggtcatg atgacgttca aaacgccaaa ctccttatcg
     1921 tgtcgccaat cgtaatcatg caggaaggcg cggccgccaa gatccaaccc gcgcgtgcgc
     1981 cagcggggtg cggcgatgaa ggcagcattg ccggctagcc cccattccgg ccgtacttgc
     2041 gaccagtctc gccctcggaa cgtggcctgt ttttcggcat caacggtatc gacttgggcc
     2101 tcaagcgtca ccaaacgctc cagacgagtg agctcaccgg cttgtttgag cgcctgggtc
     2161 aaggatgaca attgatcact ctcgatgcct gattgatcca aatccagtag ttcgatgtgg
     2221 tctgtggtgg tgtcgtgcag ggcggccacg aaccgggtgt cacgcgggat ggcaataccc
     2281 cgttcgatca gccccagtcg cacggcggcc tcattgagca actgcaccgc gacgcgagca
     2341 ttcacatcgc ctgcctgacc ggcgcaggca ccgcaatcca accccgcgcg atgcgggttg
     2401 ttggtggtgg agctgccgtg tccggcgagc aggacgatgg gcgcaaaggt gtgcgttaac
     2461 cccagcccgc ggagtacttt ttcggccaga tcgacttttt cctgcgtcga tagtgcgccg
     2521 ccatcgagtt tgaccaattg gggatgcaaa cgcgcacgct cctcaggggt aagcccgggg
     2581 gcatcaggcg gcagcgatga gcgatgccaa cccagtgaat cggccagcaa acgcggcacg
     2641 tagctcaagc ctgcggattc gacaaaggtg aagcacgagg cggcggaaag tttgaattgc
     2701 ttccagatgg cccctcggct ttgtcgggca tggcgatgtt gggcaatggc cggatcgccc
     2761 gtctcctgcg cacggtaggc ggggttgatg agcaccgggt tttgtagccg agcctctgat
     2821 tcgcccatgc ggcaataatc aatcggcagg ccgaaaaaac cggcaaaacc gatggtttcg
     2881 agccccgggc tgctggcttc gaggtggcgg cggaacacct cggaacgcac cgcaatgcaa
     2941 aacgcggcct gaacagtcgg gcgttcggct tgctgatcgg gtgcatcggc aggctggcgg
     3001 ttcaagccgg cattgatccg gcgtcgaaac gcctgctcgg cggccgccag caggatttca
     3061 tcgatccggc tggattgagc cttagtcgaa gcagtagctg tcgccgaagg gcttgcgacc
     3121 aatttggccg gatcaaacca atcgtttagc ttcgcgcgcc attgttgccg tccagccgag
     3181 ctggtttctt tgatcaccaa agcaacccag accaagcgaa tgatcagcaa atcgcgtaaa
     3241 tcgtgttgcg actcgccgtg caaaccggct tgaaagtgta gatagcggca ccagctcgcc
     3301 cagccaccga tggtttttaa taaggtgaac aggtagtcgg gcaggcgact ttcgggcaga
     3361 ttgatctggt ccagcgccca gaacaatgca tcaatggcat cgcttgggac ggcaagcaag
     3421 tgttggcgaa cctgctttaa gccgacggcg ctggcgctgc ggtcaatcaa ggtgtagcgg
     3481 cgccattggg cgaataaact ggcactggct tctttgggta agtgccagag ggcttgcccg
     3541 cgatcatagt agttggccat gaactggctg acttgctcga tgacaaattc agatactggc
     3601 ggcgcgtcgc ggcgatcgag ctcgtttgtg accaaaggca gcgcgggtgc aggttgtcga
     3661 tggacggcca gttgctgctt gatggtatca aggctaattg atgggtccag ttgctgcgcc
     3721 gcctgcgcca aatcgtcatc ggtgatttcc ccttgggcga tcttgtcggc aaaccagccg
     3781 tgatccataa atagggattc gcccacggtt tgttccagat accgacccac cgtatcaaag
     3841 ggttgatcaa tcaaaccaag ataggggttt acggcgacaa acgaatccag tggccagatg
     3901 ggcgcgatgc gcaggcaggc tttgtcaatc aggtcaacaa tgtggtttcg ttgcgcttcg
     3961 ctgcgttgta atgatgtcag ggtcgtcatg gtcgaaatcc tcgggtaaat ggtgtgtgcc
     4021 tttgtggcag ggatttacgg ctcgataccg gatgggttag gcggaatcac ggcgcaggat
     4081 tttttcggtg cgcggccaac tttctccggc cagaaacgga aggccaagcg ttcgacccag
     4141 cggtcgagat acagcccgtt attgaggtga acaaacaagg cgaattgtcg ctccgggctc
     4201 atcaaggttt tgccggggcc ttgaagccag gcaacaaaca ggaaggttat gatcgtgatc
     4261 gccatcaaca gcagatcgag cagcggcgca cgttgcggtg tcgcggccag cgtgtcggca
     4321 aagcctgtga tgaacacctc atgcaggaag acatacacgc cggtcatggc gattgccgcg
     4381 cctgcggcga ccagaatttc acgccatgtg ccatggctca atgccttgag catcaactga
     4441 gcggtcgcca aggcgatgat ggtcagcacg gcgggcaaca tcggctcctg atcgatcgat
     4501 aaacccatcg ccgccgcgat accgagggtg aacaagccgc tcacaatcac cgtggtaaac
     4561 caggccacaa cagagattgg gcggtttttg agcactttgg cctgacgcag gtgatcggtc
     4621 atgctgcccg aggacaagaa cgcatgggcc ttgtagagcg agtggccaac caagtgcagc
     4681 aaggccaagc caaacaaccc caaacccagt tcgagcatca tgaagccgag ctgagcggcg
     4741 gtagaccagg ccagtgatga tttgatcgcc gattgggtca gcatgaccaa cgagccgata
     4801 gccaaggtca tcaaaccaat cagggccaga agcaacaggg cggtaccgtc ggcggccaat
     4861 aattcgctgg tgcgcagcac aatgattgcc ccgctgtaaa cgacaccggc gtgcatgagc
     4921 gcggaaaccg gcgtgggtgc ttccatgacc tgaatcagcc agccatgaaa cggaaattgc
     4981 gcgcttttaa ggatggcggc cagcacgatc agccaagcag ccacgtgcaa cgcaatggat
     5041 aggtggtctt gcgcatgaac ataggcgctg atttggctga actccaaact accgatttga
     5101 tgaccgatca agacggtggc cgcgagcaaa agggcgtcgg cagtccggct gaaaattgat
     5161 tttttatgtg tggccatgat cgcgcgcggc cgatcggcat aaaaagaaag cagtcgatgc
     5221 aaaccaaaac cggtcgcaat gatggcgagg gtgaataatc ccaaattgcc tgatatgacg
     5281 accaacagga aaaatccgcc ggtgaccgcc agcaagcgga aaaagcgggc ctgctgggga
     5341 tcaccatcga gatactgcac gctgtagcgg gcaatcacgc taagcacaaa actaaccaga
     5401 cttgccagca ccagagtcag gccattgact tgaattgaaa gcgccaataa tcccaatccg
     5461 gcagggaggc taccggcacc gagcatcaga ttctgttgcc ttgcaaatag aaaactgacg
     5521 acagcgagca gggccaggcc aaaggccgca aaactggtcc attgaacccg ttgccgataa
     5581 gcgcgcgcat aagcagcacc ttgcttgggt ttgattacag cggccataag gagcagtgcg
     5641 ggcatggcca gcaggagaag accggttgtg tggctgaact ctatagtcat ggacggtacc
     5701 tttctcctct ttaatgaatt cggtcagtgc gtcctgctga tgtgctcagt atctctatca
     5761 ctgataggga tgtcaatctc tatcactgat agggactcga ggtgaagacg aaagggcctc
     5821 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcggaa
     5881 ttgccagctg gggcgccctc tggtaaggtt gggaagccct gcaaagtaaa ctggatggct
     5941 ttcttgccgc caaggatctg atggcgcagg ggatcaagat ctgatcaaga gacaggatga
     6001 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     6061 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     6121 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     6181 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     6241 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     6301 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     6361 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     6421 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     6481 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     6541 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     6601 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     6661 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     6721 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     6781 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgagagc tcgcttggac
     6841 tcctgttgat agatccagta atgacctcag aactccatct ggatttgttc agaacgctcg
     6901 gttgccgccg ggcgtttttt attggtgaga atccaagcac tagtcatgac caaaatccct
     6961 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     7021 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     7081 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     7141 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     7201 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     7261 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggat
//
