LOCUS       sequence-359548-        3332 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone for D1.1 cloning, contains LacZ dropout.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3332)
  AUTHORS   Goosens VJ, Walker KT, Aragon SM, Singh A, Senthivel VR, Dekker L, 
            Caro-Astorga J, Buat MLA, Song W, Lee KY, Ellis T
  TITLE     Komagataeibacter Tool Kit (KTK): A Modular Cloning System for 
            Multigene Constructs and Programmed Protein Secretion from Cellulose
            Producing Bacteria.
  JOURNAL   ACS Synth Biol. 2021 Dec 17;10(12):3422-3434. doi: 
            10.1021/acssynbio.1c00358. Epub 2021 Nov 12.
  PUBMED    34767345
REFERENCE   2  (bases 1 to 3332)
  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: 
            "10.1021/acssynbio.1c00358"; journalName: "ACS Synth Biol"; date: 
            "2021-12-17- 17"; volume: "10"; issue: "12"; pages: "3422-3434"
COMMENT     Sequence Label: KTK_009
FEATURES             Location/Qualifiers
     source          1..3332
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      239..1010
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     terminator      complement(1125..1211)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    1269..1290
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1305..1310,1311..1328,1329..1335)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1343..1359
                     /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)."
     primer_bind     1348..1370
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1367..1383
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1367..1383
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1379..1702
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    1396..1452
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(1453..1470)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1453..1469)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1462..1484)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(1678..1697)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     terminator      1736..1830
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1878..1980
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1981..2640
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2047..2066)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     oriT            2789..2897
                     /note="incP origin of transfer"
     CDS             complement(join(2908..3332,1..238))
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
ORIGIN
        1 tgactgcgta ttggaccgtc cgcaaagaac gtccgatgag cttggaaagt gttttctggc
       61 tgaccaccac ggcgttctgg tggcccatct gcgccacgag gtgatgcagc agcattgccg
      121 ccgtgggttt cctcgcaata agcccggccc acgcctcatg cgctttgcgt tccgtttgca
      181 cccagtgacc gggcttgttc ttggcttgaa tgccgatttc tctggactgc gtggccatgc
      241 ttatctccat gcggtagggg tgccgcacgg ttgcggcacc atgcgcaatc agctgcaact
      301 tttcggcagc gcgacaacaa ttatgcgttg cgtaaaagtg gcagtcaatt acagattttc
      361 tttaacctac gcaatgagct attgcggggg gtgccgcaat gagctgttgc gtacccccct
      421 tttttaagtt gttgattttt aagtctttcg catttcgccc tatatctagt tctttggtgc
      481 ccaaagaagg gcacccctgc ggggttcccc cacgccttcg gcgcggctcc ccctccggca
      541 aaaagtggcc cctccggggc ttgttgatcg actgcgcggc cttcggcctt gcccaaggtg
      601 gcgctgcccc cttggaaccc ccgcactcgc cgccgtgagg ctcggggggc aggcgggcgg
      661 gcttcgccct tcgactgccc ccactcgcat aggcttgggt cgttccaggc gcgtcaaggc
      721 caagccgctg cgcggtcgct gcgcgagcct tgacccgcct tccacttggt gtccaaccgg
      781 caagcgaagc gcgcaggccg caggccggag gcttttcccc agagaaaatt aaaaaaattg
      841 atggggcaag gccgcaggcc gcgcagttgg agccggtggg tatgtggtcg aaggctgggt
      901 agccggtggg caatccctgt ggtcaagctc gtgggcaggc gcagcctgtc catcagcttg
      961 tccagcaggg ttgtccacgg gccgagcgaa gcgagccagc cggtggccgc tcgcggccat
     1021 cgtccacata tccacgggct ggcaagggag cgcagcgacc gcgcagggcg aagcccggag
     1081 agcaagcccg tagggggggc gcgcccagct gtctagggcg gcggatttgt cctactcagg
     1141 agagcgttca ccgacaaaca acagataaaa cgaaaggccc agtctttcga ctgagccttt
     1201 cgttttattt gatgccttta attaagaaga cggcgacgac ctgagaccgg cagtgagcgc
     1261 aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     1321 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     1381 gaccatgatt acgccaagct tgcatgcctg caggtcgact ctagaggatc cccgggtacc
     1441 gagctcgaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     1501 ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc
     1561 ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggc gcctgatgcg
     1621 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca ctctcagtac
     1681 aatctgctct gatgccgcat aaggtctcaa acgccctggg tcttcactag tcttggactc
     1741 ctgttgatag atccagtaat gacctcagaa ctccatctgg atttgttcag aacgctcggt
     1801 tgccgccggg cgttttttat tggtgagaat ccaggggtcc ccaataatta cgatttaaat
     1861 tggcgaaaat gagacgttga tcggcacgta agaggttcca actttcacca taatgaaata
     1921 agatcactac cgggcgtatt ttttgagtta tcgagatttt caggagctaa ggaagctaaa
     1981 atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa
     2041 cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat
     2101 attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt
     2161 cacattcttg cccgcctgat gaatgctcat ccggaatttc gtatggcaat gaaagacggt
     2221 gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa
     2281 acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat
     2341 tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag
     2401 aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg
     2461 gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc
     2521 gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat
     2581 gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa
     2641 tttgactttt gtccttttcc gctgcataac cctgcttcgg ggtcattata gcgatttttt
     2701 cggtatatcc atcctttttc gcacgatata caggattttg ccaaagggtt cgtgtagact
     2761 ttccttggtg tatccaacgg cgtcagccgg gcaggatagg tgaagtaggc ccacccgcga
     2821 gcgggtgttc cttcttcact gtcccttatt cgcacctggc ggtgctcaac gggaatcctg
     2881 ctctgcgagg ctggccgtag gccggcccta ccggcgcggc agcgttaccc gtgtcggcgg
     2941 ctccaacggc tcgccatcgt ccagaaaaca cggctcatcg ggcatcggca ggcgctgctg
     3001 cccgcgccgt tcccattcct ccgtttcggt caaggctggc aggtctggtt ccatgcccgg
     3061 aatgccgggc tggctgggcg gctcctcgcc ggggccggtc ggtagttgct gctcgcccgg
     3121 atacagggtc gggatgcggc gcaggtcgcc atgccccaac agcgattcgt cctggtcgtc
     3181 gtgatcaacc accacggcgg cactgaacac cgacaggcgc aactggtcgc ggggctggcc
     3241 ccacgccacg cggtcattga ccacgtaggc cgacacggtg ccggggccgt tgagcttcac
     3301 gacggagatc cagcgctcgg ccaccaagtc ct
//
