LOCUS       sequence-202793-        4158 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pCK01 derivative carrying tetR under a constitutive promoter and an 
            aTc-inducible PtetO promoter for control expression of genes. 
            Contains LacI OD for translational fusions on the Ct of cloned 
            genes..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4158)
  AUTHORS   Juarez JF, Lecube-Azpeitia B, Brown SL, Johnston CD, Church GM
  TITLE     Biosensor libraries harness large classes of binding domains for 
            construction of allosteric transcriptional regulators.
  JOURNAL   Nat Commun. 2018 Aug 6;9(1):3101. doi: 10.1038/s41467-018-05525-6.
  PUBMED    30082754
REFERENCE   2  (bases 1 to 4158)
  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 
            Commun."; date: "2018-08-6"; volume: "9(1):3101. doi"; pages: " 
            10.1038/s41467-018-05525-6"
COMMENT     Sequence Label: pCKTRBS-OD
FEATURES             Location/Qualifiers
     source          1..4158
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..19,20..25,26..42,43..48,49..74)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    1..19
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    26..44
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             113..121
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             complement(578..1237)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1152..1171
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(1238..1340)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      1685..1907
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             1955..2905
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     protein_bind    3431..3452
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(3467..4093)
                     /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     3998..4017
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 tccctatcag tgatagagat tgacatccct atcagtgata gagatactga gcacatcagc
       61 aggacgcact gaccggcggc cgcgcgaatt cgagctcggt acccgggtga cctaaggagg
      121 taaataaaaa gaaaaaccac cctggcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
      181 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagat
      241 cctctagagt ggacctgcag gcatgcaagc ttgcggccgc ccgtttttat caggctctgg
      301 gaggcagaat aaatgatcat atcgtcaatt attacctcca cggggagagc ctgagcaaac
      361 tggcctcagg catttgagaa gcacacggtc acactgcttc cggtagtcaa taaaccggta
      421 aaccagcaat agacataagc ggctatttaa cgaccctgcc ctgaaccgac gaccgggtcg
      481 aatttgcttt cgaatttctg ccattcatcc gcttattatc acttattcag gcgtagcaac
      541 caggcgttta agggcaccaa taactgcctt aaaaaaatta cgccccgccc tgccactcat
      601 cgcagtactg ttgtaattca ttaagcattc tgccgacatg gaagccatca caaacggcat
      661 gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca
      721 tggtgaaaac gggggcgaag aagttgtcca tattggccac gtttaaatca aaactggtga
      781 aactcaccca gggattggct gagacgaaaa acatattctc aataaaccct ttagggaaat
      841 aggccaggtt ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga aactgccgga
      901 aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc agtttgctca tggaaaacgg
      961 tgtaacaagg gtgaacacta tcccatatca ccagctcacc gtctttcatt gccatacgaa
     1021 attccggatg agcattcatc aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt
     1081 gcttattttt ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg gtctggttat
     1141 aggtacattg agcaactgac tgaaatgcct caaaatgttc tttacgatgc cattgggata
     1201 tatcaacggt ggtatatcca gtgatttttt tctccatttt agcttcctta gctcctgaaa
     1261 atctcgataa ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg
     1321 aacctcttac gtgccgatca acgtctcatt ttcgccaaaa gttggcccag ggcttcccgg
     1381 tatcaacagg gacaccagga tttatttatt ctgcgaagtg atcttccgtc acaggtattt
     1441 attcggcgcc tgtagtgcca tttaccccca ttcactgcca gagccgtgag cgcagcgaac
     1501 tgaatgtcac gaaaaagaca gcgacttagg tgcctgatgg tcggagacaa aaggaatatt
     1561 cagcgatttg cccgagcttg cgagggtgct acttaagcct ttagggtttt aaggtctgtt
     1621 ttgtagagga gcaaacagcg tttgcgacat ccttttgtaa tactgcggaa ctgactaaag
     1681 tagtgagtta tacacagggc tgggatctat tctttttatc tttttttatt ctttctttat
     1741 tctataaatt ataaccactt gaatataaac aaaaaaaaca cacaaaggtc tagcggaatt
     1801 tacagagggt ctagcagaat ttacaagttt tccagcaaag gtctagcaga atttacagat
     1861 acccacaact caaaggaaaa ggactagtaa ttatcattga ctagcccatc tcaattggta
     1921 tagtgattaa aatcacctag accaattgag atgtatgtct gaattagttg ttttcaaagc
     1981 aaatgaacta gcgattagtc gctatgactt aacggagcat gaaaccaagc taattttatg
     2041 ctgtgtggca ctactcaacc ccacgattga aaaccctaca aggaaagaac ggacggtatc
     2101 gttcacttat aaccaatacg ctcagatgat gaacatcagt agggaaaatg cttatggtgt
     2161 attagctaaa gcaaccagag agctgatgac gagaactgtg gaaatcagga atcctttggt
     2221 taaaggcttt gagattttcc agtggacaaa ctatgccaag ttctcaagcg aaaaattaga
     2281 attagttttt agtgaagaga tattgcctta tcttttccag ttaaaaaaat tcataaaata
     2341 taatctggaa catgttaagt cttttgaaaa caaatactct atgaggattt atgagtggtt
     2401 attaaaagaa ctaacacaaa agaaaactca caaggcaaat atagagatta gccttgatga
     2461 atttaagttc atgttaatgc ttgaaaataa ctaccatgag tttaaaaggc ttaaccaatg
     2521 ggttttgaaa ccaataagta aagatttaaa cacttacagc aatatgaaat tggtggttga
     2581 taagcgaggc cgcccgactg atacgttgat tttccaagtt gaactagata gacaaatgga
     2641 tctcgtaacc gaacttgaga acaaccagat aaaaatgaat ggtgacaaaa taccaacaac
     2701 cattacatca gattcctacc tacataacgg actaagaaaa acactacacg atgctttaac
     2761 tgcaaaaatt cagctcacca gttttgaggc aaaatttttg agtgacatgc aaagtaagta
     2821 tgatctcaat ggttcgttct catggctcac gcaaaaacaa cgaaccacac tagagaacat
     2881 actggctaaa tacggaagga tctgaggttc ttatggctct tgtatctatc agtgaagcat
     2941 caagactaac aaacaaaagt agaacaactg ttcaccgtta catatcaaag ggaaaactgt
     3001 ccatatgcac agatgaaaac ggtgtaaaaa agatagatac atcagagctt ttacgagttt
     3061 ttggtgcatt taaagctgtt caccatgaac agatcgacaa tgtaacagat gaacagcatg
     3121 taacacctaa tagaacaggt gaaaccagta aaacaaagca actagaacat gaaattgaac
     3181 acctgagaca acttgttaca gctcaacagt cacacataga cagcctgaaa caggcgatgc
     3241 tgcttatcga atcaaagctg ccgacaacac gggagccagt gacgcctccc gtggggaaaa
     3301 aatcatggca attctggaag aaatagcgcc caatacgcaa accgcctctc cccgcgcgtt
     3361 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     3421 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttaa gacccacttt
     3481 cacatttaag ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg
     3541 gctctgcacc ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat
     3601 cagtagtagg tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc
     3661 aacctaaagt aaaatgcccc acagcgctga gtgcatataa tgcattctct agtgaaaaac
     3721 cttgttggca taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc
     3781 gtgtacctaa atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt
     3841 tagcgttatt acgtaaaaaa tcttgccagc tttccccttc taaagggcaa aagtgagtat
     3901 ggtgcctatc taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat
     3961 gccaatacaa tgtaggctgc tctacaccta gcttctgggc gagtttacgg gttgttaaac
     4021 cttcgattcc gacctcatta agcagctcta atgcgctgtt aatcacttta cttttatcta
     4081 atctagacat cattaattcc taatttttgg gtctactcta tcattgatag cgtgccttta
     4141 ccactcccta tcatcgtt
//
