LOCUS       40924_26181        5093 bp DNA     circular SYN 13-MAY-2021
DEFINITION  Tetracycline biosensor. Low copy plasmid that expresses GFP 
            proportional to tetracycline concentration..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5093)
  AUTHORS   Rogers JK, Guzman CD, Taylor ND, Raman S, Anderson K, Church GM
  TITLE     Synthetic biosensors for precise gene control and real-time 
            monitoring of metabolites.
  JOURNAL   Nucleic Acids Res. 2015 Jul 7. pii: gkv616.
  PUBMED    26152303
REFERENCE   2  (bases 1 to 5093)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5093)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2015 Jul 7. pii: gkv616."
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5093
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             59..916
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(1076..2023)
                     /codon_start=1
                     /label=Rep101
                     /note="RepA protein needed for replication with the pSC101 
                     origin"
                     /translation="MSKLVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKHGRI"
     rep_origin      complement(2071..2293)
                     /direction=LEFT
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     primer_bind     complement(2785..2802)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse
                     primer"
     terminator      2955..3041
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     promoter        3203..3275
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator
                     sites (Lutz and Bujard, 1997)"
     CDS             3313..4026
                     /codon_start=1
                     /label=superfolder GFP
                     /note="GFP variant that folds robustly even when fused to
                     poorly folded proteins (Pedelacq et al., 2006)"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     protein_bind    complement(4055..4076)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     RBS             4319..4327
                     /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             4335..4955
                     /codon_start=1
                     /label=TetR
                     /note="tetracycline repressor TetR"
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     terminator      5012..5059
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
       61 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
      121 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
      181 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
      241 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
      301 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
      361 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
      421 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
      481 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
      541 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
      601 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
      661 ccggcaacaa ttgatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
      721 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggctctcg
      781 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
      841 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
      901 actgattaag cattggtaac tgtcatacca agtttactca tatatacttt agattgattt
      961 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     1021 caaaatccct taacgtggag ttttcgttcc actgagcgtc agaccccgaa cctcagatcc
     1081 ttccgtgttt agccagtatg ttctctagtg tggttcgttg tttttgcgtg agccatgaga
     1141 acgaaccatt gagatcatac ttactttgca tgtcactcaa aaattttgcc tcaaaactgg
     1201 tgagctgaat ttttgcagtt aaagcatcgt gtagtgtttt tcttagtccg ttatgtaggt
     1261 aggaatctga tgtaatggtt gttggtattt tgtcaccatt catttttatc tggttgttct
     1321 caagttcggt tacgagatcc atttgtctat ctagttcaac ttggaaaatc aacgtatcag
     1381 tcgggcggcc tcgcttatca accaccaatt tcatattgct gtaagtgttt aaatctttac
     1441 ttattggttt caaaacccat tggttaagcc ttttaaactc atggtagtta ttttcaagca
     1501 ttaacatgaa cttaaattca tcaaggctaa tctctatatt tgccttgtga gttttctttt
     1561 gtgttagttc ttttaataac cactcataaa tcctcataga gtatttgttt tcaaaagact
     1621 taacatgttc cagattatat tttatgaatt tttttaactg gaaaagataa ggcaatatct
     1681 cttcactaaa aactaattct aatttttcgc ttgagaactt ggcatagttt gtccactgga
     1741 aaatctcaaa gcctttaacc aaaggattcc tgatttccac agttctcgtc atcagctctc
     1801 tggttgcttt agctaataca ccataagcat tttccctact gatgttcatc atctgagcgt
     1861 attggttata agtgaacgat accgtccgtt ctttccttgt agggttttca atcgtggggt
     1921 tgagtagtgc cacacagcat aaaattagct tggtttcatg ctccgttaag tcatagcgac
     1981 taatcgctag ttcatttgct ttgaaaacaa ctaatttaga catacatctc aattggtcta
     2041 ggtgatttta atcactatac caattgagat gggctagtca atgataatta ctagtccttt
     2101 tcctttgagt tgtgggtatc tgtaaattct gctagacctt tgctggaaaa cttgtaaatt
     2161 ctgctagacc ctctgtaaat tccgctagac ctttgtgtgt tttttttgtt tatattcaag
     2221 tggttataat ttatagaata aagaaagaat aaaaaaagat aaaaagaata gatcccagcc
     2281 ctgtgtataa ctcactactt tagtcagttc cgcagtatta caaaaggatg tcgcaaacgc
     2341 tgtttgctcc tctacaaaac agaccttaaa accctaaagg cttaagtagc accctcgcaa
     2401 gctcgggcaa atcgctgaat attccttttg tctccgacca tcaggcacct gagtcgctgt
     2461 ctttttcgtg acattcagtt cgctgcgctc acggctctgg cagtgaatgg gggtaaatgg
     2521 cactacaggc gccttttatg gattcatgca aggaaactac ccataataca agaaaagccc
     2581 gtcacgggct tctcagggcg ttttatggcg ggtctgctat gtggtgctat ctgacttttt
     2641 gctgttcagc agttcctgcc ctctgatttt ccagtctgac cacttcggat tatcccgtga
     2701 caggtcattc agactggcta atgcacccag taaggcagcg gtatcatcaa ctggctgttt
     2761 tggcggatga gagaagattt tcagcctgat acagattaaa tcagaacgca gaagcggtct
     2821 gataaaacag aatttgcctg gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa
     2881 ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg tctccccatg cgagagtagg
     2941 gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta
     3001 tctgttgttt gtcggtgaac gctctcctga gtaggacaaa tccgccggga gcggatttga
     3061 acgttgcgaa gcaacggccc ggagggtggc gggcaggacg cccgccataa actgccaggc
     3121 atcaaattaa gcagaaggcc atccgtcagg atggcctttt tgcgtttcta caaactctgc
     3181 tagcaagtaa ggccgactcg agtccctatc agtgatagag attgacatcc ctatcagtga
     3241 tagagatact gagcacatca gcaggacgca tgaccgaatt cattaaattt aactttaaga
     3301 aggagatata atatgcgtaa aggtgaagaa ctgttcaccg gtgttgttcc gatcctggtt
     3361 gaactggacg gtgacgttaa cggtcacaaa ttctctgttc gtggtgaagg tgaaggtgac
     3421 gctaccaacg gtaaactgac cctgaaattc atctgcacca ccggtaaact gccggttccg
     3481 tggccgaccc tggttaccac cctgacctac ggtgttcagt gcttcgctcg ttacccggac
     3541 cacatgaaac agcacgactt cttcaaatct gctatgccgg aaggttacgt tcaggaacgt
     3601 accatctctt tcaaagacga cggtacctac aaaacccgtg ctgaagttaa attcgaaggt
     3661 gacaccctgg ttaaccgtat cgaactgaaa ggtatcgact tcaaagaaga cggtaacatc
     3721 ctgggtcaca aactggaata caacttcaac tctcacaacg tttacatcac cgctgacaaa
     3781 cagaaaaacg gtatcaaagc taacttcaaa atccgtcaca acgttgaaga cggttctgtt
     3841 cagctggctg accactacca gcagaacacc ccgatcggtg acggtccggt tctgttgccg
     3901 gacaaccact acctgtctac ccagtctgtt ctgtctaaag acccgaacga aaaacgtgac
     3961 cacatggttc tgctggaatt cgttaccgct gctggtatca cccacggtat ggacgaactg
     4021 tacaaagatg catgccagtt ctagcataac cctaatgagt gagctaactt acattaattg
     4081 cgttgcgcct taattaacgg cactcctcag caaatataat gaccctcttg ataacccaag
     4141 agggcatttt ttaatgccca tggcgtttac cacagctaac accacgtcgt ccctatctgc
     4201 tgccctaggt ctatgagtgg ttgctggata actttacggg catgcataag gctcgtaggc
     4261 tatattcagg gagaccacaa cggtttccct ctacaaataa ttttgtttaa ctttgaaata
     4321 aggaggtaat acaaatgtct cgtttagata aaagtaaagt gattaacagc gcattagagc
     4381 tgcttaatga ggtcggaatc gaaggtttaa caacccgtaa actcgcccag aagctaggtg
     4441 tagagcagcc tacattgtat tggcatgtaa aaaataagcg ggctttgctc gacgccttag
     4501 ccattgagat gttagatagg caccatactc acttttgccc tttagaaggg gaaagctggc
     4561 aagatttttt acgtaataac gctaaaagtt ttagatgtgc tttactaagt catcgcgatg
     4621 gagcaaaagt acatttaggt acacggccta cagaaaaaca gtatgaaact ctcgaaaatc
     4681 aattagcctt tttatgccaa caaggttttt cactagagaa tgcattatat gcactcagcg
     4741 ctgtggggca ttttacttta ggttgcgtat tggaagatca agagcatcaa gtcgctaaag
     4801 aagaaaggga aacacctact actgatagta tgccgccatt attacgacaa gctatcgaat
     4861 tatttgatca ccaaggtgca gagccagcct tcttattcgg ccttgaattg atcatatgcg
     4921 gattagaaaa acaacttaaa tgtgaaagtg ggtcttaata agcgactaaa aaattgaatg
     4981 taggaaacca acatgccagt tcgagcaata actagcataa ccccttgggg cctctaaacg
     5041 ggtcttgagg ggttttttgc tgaaacctca ggcgagaagc acacggtcac act
//
