LOCUS       sequence-267950-        4520 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing synthetic overlapping gene, ilvA-ccdB .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4520)
  AUTHORS   Blazejewski T, Ho HI, Wang HH
  TITLE     Synthetic sequence entanglement augments stability and containment 
            of genetic information in cells.
  JOURNAL   Science. 2019 Aug 9;365(6453):595-598. doi: 10.1126/science.aav5477.
  PUBMED    31395784
REFERENCE   2  (bases 1 to 4520)
  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"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2019-08-9- 9"; volume: "365"; issue: 
            "6453"; pages: "595-598"
COMMENT     Sequence Label: pH9-ilvA-ccdBwt (DE25)
FEATURES             Location/Qualifiers
     source          1..4520
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      1569..1640
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1656..1683
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(1711..1728)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1845..2390)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(1925..1944)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2504..2598)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2629..3420)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVITEVSTQLSEVVGAIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     CDS             complement(3788..4414)
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     4319..4338
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        4430..4485
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    4466..4484
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             4503..4514
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
ORIGIN
        1 atggctgact cgcaacccct gtccggtgct ccggaaggtg ccgaatattt aagagcagtg
       61 ctgcgcgcgc cggtttacga ggcggcgcag gttacgccgc tacaaaaaat ggaaaaactg
      121 tcgtcgcgtc ttgataacgt cattctggtg aagcgcgaag atcgccagcc agtgcacagc
      181 tttaagctgc gcggcgcata cgccatgatg gcgggcctga cggaagaaca gaaagcgcac
      241 ggcgtgatca ctgcttctgc gggtaaccac gcgcagggcg tcgcgttttc ttctgcgcgg
      301 ttaggcgtga aggccctgat cgttatgcca accgccaccg ccgacatcaa agtcgacgcg
      361 gtgcgcggct tcggcggcga agtgctgctc cacggcgcga actttgatga agcgaaagcc
      421 aaagcgatcg aactgtcaca gcagcagggg ttcacctggg tgccgccgtt cgaccatccg
      481 atggtgattg ccgggcaagg cacgctggcg ctggaactgc tccagcagga cgcccatctc
      541 gaccgcgtat ttgtgccagt cggcggcggc ggtctggctg ctggcgtggc ggtgctgatc
      601 aaacaactga tgccgcaaat caaagtgatc gccgtagaag cggaagactc cgcctgcctg
      661 aaagcagcgc tggatgcggg tcatccggtt gatctgccgc gcgtagggct atttgctgaa
      721 ggcgtagcgg taaaacgcat cggtgacgaa accttccgtt tatgccagga gtatctcgac
      781 gacatcatca ccgtcgatag cgatgcgatc tgtgcggcga tgaaggattt attcgaagat
      841 gtgcgcgcgg tggcggaacc ctctggcgcg ctggcgctgg cgggaatgaa aaaatatatc
      901 gccctgcaca acattcgcgg cgaacggctg gcgcatattc tttccggtgc caacgtgaac
      961 ttccacggcc tgcgctacgt ctcagaacgc tgcgaactgg gcgaacagcg tgaagcgttg
     1021 ttggcggtga ccattaatgc agttcaaggt gtatacctac aagcgggagt cgaggtatcg
     1081 gctcttcgtg gacgtacaga gcgacattat agatacgccg gggaggagga tggtcatacc
     1141 tttagcgtca gccaggttgc tgagcgacaa agtaagcaga gaattatatc ccgtagtaca
     1201 cataggggac gaaagctgga ggatgatgac gacggacatg gcgtccgtac ccgtgtccgt
     1261 tataggagag gaggtcgcgg atctaagtca cagagagaac gatataaaaa acgcgataaa
     1321 cctgatgttc tgggggatat gattcgcttc ctcaacacgc tgggtacgta ctggaacatt
     1381 tctttgttcc actatcgcag ccatggcacc gactacgggc gcgtactggc ggcgttcgaa
     1441 cttggcgacc atgaaccgga tttcgaaacc cggctgaatg agctgggcta cgattgccac
     1501 gacgaaacca ataacccggc gttcaggttc tttttggcgg gttagctcga gtaaggatct
     1561 ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt
     1621 gtttgtcggt gaacgctctc tactagagtc acactggctc accttcgggt gggcctttct
     1681 gcgtttatac ctagggatat attccgcttc ctcgctcact gactcgctac gctcggtcgt
     1741 tcgactgcgg cgagcggaaa tggcttacga acggggcgga gatttcctgg aagatgccag
     1801 gaagatactt aacagggaag tgagagggcc gcggcaaagc cgtttttcca taggctccgc
     1861 ccccctgaca agcatcacga aatctgacgc tcaaatcagt ggtggcgaaa cccgacagga
     1921 ctataaagat accaggcgtt tccccctggc ggctccctcg tgcgctctcc tgttcctgcc
     1981 tttcggttta ccggtgtcat tccgctgtta tggccgcgtt tgtctcattc cacgcctgac
     2041 actcagttcc gggtaggcag ttcgctccaa gctggactgt atgcacgaac cccccgttca
     2101 gtccgacagc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg aaagacatgc
     2161 aaaagcacaa ctggcagcag ccactggtaa ttgatttaga ggagttagtc ttgaagtcat
     2221 gcgccggtta aggctaaact gaaaggacaa gttttggtga ctgcgctcct ccaagccagt
     2281 tacctcggtt caaagagttg gtagctcaga gaaccttcga aaaaccgccc tgcaaggcgg
     2341 ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa acgatctcaa gaagatcatc
     2401 ttattaatca gataaaatat ttctagattt cagtgcaatt tatctcttca aatgtagcac
     2461 ctgaagtcag ccccatacga tataagttgt tactagtgct tggattctca ccaataaaaa
     2521 acgcccggcg gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg
     2581 gatctatcaa caggagtcca agcgagctct cgaaccccag agtcccgctt atttgccgac
     2641 taccttggtg atctcgcctt tcacgtagtg gacaaattct tccaactgat ctgcgcgcga
     2701 ggccaagcga tcttcttctt gtccaagata agcctgtcta gcttcaagta tgacgggctg
     2761 atactgggcc ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt
     2821 gccggttact gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc
     2881 agcccagtcg ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc
     2941 ctgttcagga accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg
     3001 ttctcttgct tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat
     3061 acctgcaaga atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata
     3121 acgccacgga atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc
     3181 gctctctcca ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt
     3241 ttcatcaagc cttacggtca ccgtaactag caaatcaata tcactgtgtg gcttcaggcc
     3301 gccatccact gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc
     3361 gatggcgcca actacctctg atagttgagt cgatacttcg gtgatcaccg cttccctcat
     3421 aatgtttaac tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctccataa
     3481 catcaaacat cgacccatgg cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg
     3541 taccccaaaa aaacagtcat agcaagccat gaaaaccgcc actgcaccgt taccaccgct
     3601 gcgttcggtc aaggttctgg accagttgcg tgagcgcatg cgaaacgatc ctcatcctgt
     3661 ctcttgatca gatcttgatc ccctgcgcca tcagatcctt ggcggcaaga aagccatcca
     3721 gtttactttg cagggcttcc caaccttacc agagggcgcc ccagctggca attccgacgt
     3781 cgacgtctta agacccactt tcacatttaa gttgtttttc taatccgcat atgatcaatt
     3841 caaggccgaa taagaaggct ggctctgcac cttggtgatc aaataattcg atagcttgtc
     3901 gtaataatgg cggcatacta tcagtagtag gtgtttccct ttcttcttta gcgacttgat
     3961 gctcttgatc ttccaatacg caacctaaag taaaatgccc cacagcgctg agtgcatata
     4021 atgcattctc tagtgaaaaa ccttgttggc ataaaaaggc taattgattt tcgagagttt
     4081 catactgttt ttctgtaggc cgtgtaccta aatgtacttt tgctccatcg cgatgactta
     4141 gtaaagcaca tctaaaactt ttagcgttat tacgtaaaaa atcttgccag ctttcccctt
     4201 ctaaagggca aaagtgagta tggtgcctat ctaacatctc aatggctaag gcgtcgagca
     4261 aagcccgctt attttttaca tgccaataca atgtaggctg ctctacacct agcttctggg
     4321 cgagtttacg ggttgttaaa ccttcgattc ctacctcatt aagcagctct aatgcgctgt
     4381 taatcacttt acttttatct aatctagaca tcattaattc ctaatttttg ttgacactct
     4441 atcgttgata gagttatttt accactccct atcagtgata gagaaaagaa ttcaaaagat
     4501 ctaaagagga gaaaggatct
//
