LOCUS       sequence-378902-        8091 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Level 2 MoClo plasmid for A. baumannii.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8091)
  AUTHORS   Brychcy M, Kokodynski A, Lloyd D, Godoy VG
  TITLE     AspFlex: Molecular Tools to Study Gene Expression and Regulation in 
            Acinetobacter baumannii.
  JOURNAL   ACS Synth Biol. 2023 Aug 16. doi: 10.1021/acssynbio.3c00167.
  PUBMED    37587063
REFERENCE   2  (bases 1 to 8091)
  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: "ACS Synth 
            Biol. 2023 Aug 16. doi: 10.1021/acssynbio.3c00167."
COMMENT     Sequence Label: pMB2b
FEATURES             Location/Qualifiers
     source          1..8091
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    123..144
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(159..164,165..182,183..189)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    197..213
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             231..242
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             249..926
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      971..1042
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1058..1085
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      5046..5103
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(5127..5144)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5298..5886)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5378..5397)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(6069..6163)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             6267..7061
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     terminator      7079..7165
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(7182..7841)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     7756..7775
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     8003..8021
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(8025..8068)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    join(8071..8091,1)
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 ggtctcacta tatctagaga cgcaatacgc aaaccgcctc tccccgcgcg ttggccgatt
       61 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
      121 attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct
      181 cgtatgttgt gtggaattgt gagcggataa caatttcaca catactagag aaagaggaga
      241 aatactagat ggcttcctcc gaagacgtta tcaaagagtt catgcgtttc aaagttcgta
      301 tggaaggttc cgttaacggt cacgagttcg aaatcgaagg tgaaggtgaa ggtcgtccgt
      361 acgaaggtac ccagaccgct aaactgaaag ttaccaaagg tggtccgctg ccgttcgctt
      421 gggacatcct gtccccgcag ttccagtacg gttccaaagc ttacgttaaa cacccggctg
      481 acatcccgga ctacctgaaa ctgtccttcc cggaaggttt caaatgggaa cgtgttatga
      541 acttcgaaga cggtggtgtt gttaccgtta cccaggactc ctccctgcaa gacggtgagt
      601 tcatctacaa agttaaactg cgtggtacca acttcccgtc cgacggtccg gttatgcaga
      661 aaaaaaccat gggttgggaa gcttccaccg aacgtatgta cccggaagac ggtgctctga
      721 aaggtgaaat caaaatgcgt ctgaaactga aagacggtgg tcactacgac gctgaagtta
      781 aaaccaccta catggctaaa aaaccggttc agctgccggg tgcttacaaa accgacatca
      841 aactggacat cacctcccac aacgaagact acaccatcgt tgaacagtac gaacgtgctg
      901 aaggtcgtca ctccaccggt gcttaataac gctgatagtg ctagtgtaga tcgctactag
      961 agccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     1021 ttgtttgtcg gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt
     1081 ctgcgtttat acgtctcaga aggtacagag accctgcagt tgcaagacaa tatcgaccgc
     1141 ttgcagcttg ataaagtgat caaagaggga cgagcagcta cagcactgct aagggagatc
     1201 ggagagccac tgccgccacc agtgaaagtg aaaacaggtt ggtttagtgc tgattattac
     1261 ggctttgagg acatgctcac acgttgggat aatgatcaat ctagcctttc catttatcac
     1321 agagagataa aagaggagca acaggaacag caacgaaaac aacagcagat agaaagagaa
     1381 actaaacaaa aacaggaaga tgacaaaaag aaacggcagg actatgaaag atatttatca
     1441 aaatacggct atgtcaaaga tgctgagtat gaccatatca gcgaccatgt tgcttcagac
     1501 atttccatgt ttacacgtct tcaggcgcag gcatgggcaa gtaacgatat gcaagagtac
     1561 ataaggtgtt caaagcaaaa atacgagcag atcagcgaca gaatagccta tgagcgtgac
     1621 ttgaaaaaaa tcgggcaatt gagccgtatc ctggataaag acagacaggc acttgctgat
     1681 attttaccgc agttgatgcc ttactatgag cagatgcaac aggctgttaa taaacggaaa
     1741 attttacttg agaatgagcg acagccatca tttaaaccac gttatgacca agaacagcct
     1801 aagccgaaaa aagacaatga cctaactttt taatgtgtta caatagcatt caaatgtaac
     1861 acattggagt gaaaaacatg acagaagcga cttttacctt tcgggtagac catgacctaa
     1921 agcaagaatt ttcaagcctt gctaagactg ttgatcggtc gggggcgcag ctaatccgtg
     1981 acttcatgcg tgactttgtt aagaagcaac aggaagccgc agactatgac aaatggttca
     2041 agcagcaggt acagatcggc ttaaatgaag ccaatgcagg caagttgatt ccacatgagg
     2101 acgtaaaggc agagtttgcc gctagacgtg ctgcaacatt ggcgaaactg gcagcgaaga
     2161 atgaagattg agtggactga aacagctcgc caagatcgta gaaatatcta tgattatctt
     2221 gaagaacgca accctatagc agctattgaa attgatgatt taattgaaga aaagacagat
     2281 ttacttgttg ataatcgact gatggggcgc acaggcagac agaaagatac tagggagtta
     2341 gtgatacatc cgcattatgt ggttgtatat gacatcactg atataatacg gatactcaga
     2401 gtgctacaca catcgcagga gtggtcatga cttactcatg tactttggat tatttagtgt
     2461 tataaaatcc tgatttataa attttttttg ttaaaaaaga taaaagcccc ttgcaattgc
     2521 ttggggcttt accgtaattt atggggtaca gatcttcgat actgacatat cggcaatcga
     2581 aagcattaag gtttgacgac cgctaatgat ttcaccacag gggcttaatg tacctgtctt
     2641 aaattctaag gttttaactc gctttgtcaa gcatagaccc caaaaattta gccaatgtct
     2701 gtaactcaat ctgtccatgt gtgggtgatg aggtacagtg acgctagcac acatcggaaa
     2761 aacgctatta ctaggggaac tgaacagagt agcggacgca atgagtagtc atttaattgg
     2821 cggttatgag cgtgttcagg cggtgctatc aatcgtaatc ataacagtgg cagcttgata
     2881 cagtgatgtc atccctgatg cgaaagcgac cgaccgacgg tacatcgaat gggaatactt
     2941 tagggtgatt tttaagaatc gctctagggt gagtatttcc cattcagctc tgctccctcc
     3001 ctctggtact ttaatcaaaa gcactactaa acatatgttt ttaaataaaa aatattgata
     3061 tagagataat attagtaaga ataattaaac aattgaatat agataaatca ttgttaaata
     3121 aagattaatt attaaaatga atgtatactt atatataaat caatgattta aaatatttga
     3181 taaagaaaac ttttcaaaaa aaatataatt gagattgtgt catttcggtc aattcttaat
     3241 atgttccacg caagttttag ctatggtgct aaacagaaat ttgctgaaaa agaacttttc
     3301 actgaactgg ttaaaatgta agcagcctga gagccgccaa aaattttaaa aacaaaccgc
     3361 cttaatcatc ttcaaaaaat acctctaaaa cctcaccatt tgcgttttaa gacccatatt
     3421 tcatcctgcc cttatgttcc catgctgata gctataaagt gtctgtaatc gcttcctatg
     3481 acgttctagg ctgttgataa cttttggaac aacgcaaaat gttaaaatcc gatcattttt
     3541 taacctagtt attttcgtta cagggtaact ttggtagtat tatttcaata tttagttgtt
     3601 acaggataac aaaatatatg ttacagggta attaatttag cgttacagga taactatagt
     3661 tacagggtaa cgttaaatag ttatcctgta atttaaaaat agttgcagga tagcataaat
     3721 ttttacaaat aaattttata aaaagttatc ctgtaactaa aaacaatgtt atcctgtaac
     3781 tatgatatac tatatgggag ttaaagacat gaaagacccg aacgatcaga aaacacagga
     3841 catgctaaaa gagccgccaa aaaccaatgc cgaacgacaa aaagcatatc gtgaaaaacg
     3901 caaaagcctt gatagtcaac gtttggaggt gtttatagat aagggtgtgt cggatatgct
     3961 tgcggacatg gtgggcgcag caggggagag ccaaaaagcc attttgaccg cattgattga
     4021 gaaagagtat aagcggctgt atgcagtgaa aaatagcaag ctaaaaaagt acattgattc
     4081 aaaaagtaat gcagataaaa aagaaaccct cgatttatga gggtttttta taactactaa
     4141 tgcctatgaa aataagtgct tatcattttc tctaatgtac ccaataaaat cggggttttg
     4201 ttccacaagt ttcttgatgt cattatcatg tttcttcagt aagtaagccc cgatcaggat
     4261 cttgcgtttt gcatcgtctg accttgcctg tttttgctct cgacctttca ccacagcgtc
     4321 agctagatcc tttcttgcct tttcctgacg ttccaatttt tccttttctt catcgagcat
     4381 ttgacccaat tccatagctg tttttttgac catttttaat gcctgtgtct caacctattt
     4441 tccgaagtat gaaccatcat caatgtgaaa agtgttctac ttcaaaaact cgacttacat
     4501 caagatccga gcgcagcgag caaaatcaaa aaacaaagtc aaagccttat tgctcttgct
     4561 tttgctttta gcctcgcaga gttcccgaag ggcgcactta cgcaaaattt ttgctacgcc
     4621 aaattttgca agtacggtca gggaaacccc gacaccccaa ccgcccaaaa cttggggcgg
     4681 ctgttaataa acaggtgatg aaaatggcaa tttaccattg tgaaatgcag aacatttcga
     4741 ggtcagatgg tcgctcaatc gtggcatgtg cagcataccg agcaggcgaa aaattgtact
     4801 gtgatacgta cggaaaagag caggactaca ccaaaaaaac aggcattgaa tacacccaaa
     4861 tttttgcccc aactggggca agtcctgaca tgttagatcg tcaaacccta tggaatcggg
     4921 tagagcaatc cgaactaaaa aagaacggtg acatcaaaca ggaggcaaga ttagcaaagg
     4981 aagtagagat agcattgccg catgaactgg ataagacaca gcgtcaggca acttgttaac
     5041 tgcagtccgg caaaaaaggg caaggtgtca ccaccctgcc ctttttcttt aaaaccgaaa
     5101 agattacttc gcgttatgca ggcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     5161 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5221 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5281 gccgcgttgc tggcgttttt ccacaggctc cgcccccctg acgagcatca caaaaatcga
     5341 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5401 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     5461 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     5521 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     5581 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     5641 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     5701 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     5761 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     5821 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     5881 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     5941 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     6001 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagctcg
     6061 aggcttggat tctcaccaat aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc
     6121 agatggagtt ctgaggtcat tactggatct atcaacagga gtccaagcga gctcgatatt
     6181 ctcgagaagc tggggatccg tttgattttt aatggataat gtgatataat ctttaaatac
     6241 tgtagaaaag aggaaggaaa taataaatgg ctaaaatgag aatatcaccg gaattgaaaa
     6301 aactgatcga aaaataccgc tgcgtaaaag atacggaagg aatgtctcct gctaaggtat
     6361 ataagctggt gggagaaaat gaaaacctat atttaaaaat gacggacagc cggtataaag
     6421 ggaccaccta tgatgtggaa cgggaaaagg acatgatgct atggctggaa ggaaagctgc
     6481 ctgttccaaa ggtcctgcac tttgaacggc atgatggctg gagcaatctg ctcatgagtg
     6541 aggccgatgg cgtcctttgc tcggaagagt atgaagatga acaaagccct gaaaagatta
     6601 tcgagctgta tgcggagtgc atcaggctct ttcactccat cgacatatcg gattgtccct
     6661 atacgaatag cttagacagc cgcttagccg aattggatta cttactgaat aacgatctgg
     6721 ccgatgtgga ttgcgaaaac tgggaagaag acactccatt taaagatccg cgcgagctgt
     6781 atgatttttt aaagacggaa aagcccgaag aggaacttgt cttttcccac ggcgacctgg
     6841 gagacagcaa catctttgtg aaagatggca aagtaagtgg ctttattgat cttgggagaa
     6901 gcggcagggc ggacaagtgg tatgacattg ccttctgcgt ccggtcgatc agggaggata
     6961 tcggggaaga acagtatgtc gagctatttt ttgacttact ggggatcaag cctgattggg
     7021 agaaaataaa atattatatt ttactggatg aattgtttta ggacgtcgcc ggcggcatca
     7081 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt
     7141 gaacgctctc ctgagtagga caaatcctcg agaatatcaa attacgcccc gccctgccac
     7201 tcatcgcagt actgttgtaa ttcattaagc attctgccga catggaagcc atcacaaacg
     7261 gcatgatgaa cctgaatcgc cagcggcatc agcaccttgt cgccttgcgt ataatatttg
     7321 cccatggtga aaacgggggc gaagaagttg tccatattgg ccacgtttaa atcaaaactg
     7381 gtgaaactca cccagggatt ggctgacacg aaaaacatat tctcaataaa ccctttaggg
     7441 aaataggcca ggttttcacc gtaacacgcc acatcttgcg aatatatgtg tagaaactgc
     7501 cggaaatcgt cgtggtattc actccagagc gatgaaaacg tttcagtttg ctcatggaaa
     7561 acggtgtaac aagggtgaac actatcccat atcaccagct caccgtcttt cattgccata
     7621 cgaaattccg gatgagcatt catcaggcgg gcaagaatgt gaataaaggc cggataaaac
     7681 ttgtgcttat ttttctttac ggtctttaaa aaggccgtaa tatccagctg aacggtctgg
     7741 ttataggtac attgagcaac tgactgaaat gcctcaaaat gttctttacg atgccattgg
     7801 gatatatcaa cggtggtata tccagtgatt tttttctcca ttttagcttc cttagctcct
     7861 gaaaatctcg ataactcaaa aaatacgccc ggtagtgatc ttatttcatt atggtgaaag
     7921 ttggaacctc ttacgtgccc gatcaactcg agtgccacct gacgtctaag aaaccattat
     7981 tatcatgaca ttaacctata aaaataggcg tatcacgagg cagaatttca gataaaaaaa
     8041 atccttagct ttcgctaagg atgatttctg gaattcgcgg ccgcttctag a
//
