LOCUS       sequence-274915-        3831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Ribosome Binding Site (RBS) variant using GFP as a reporter 
            gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3831)
  AUTHORS   Teh MY, Ooi KH, Danny Teo SX, Bin Mansoor ME, Shaun Lim WZ, Tan MH
  TITLE     An Expanded Synthetic Biology Toolkit for Gene Expression Control in
            Acetobacteraceae.
  JOURNAL   ACS Synth Biol. 2019 Apr 19;8(4):708-723. doi: 
            10.1021/acssynbio.8b00168. Epub 2019 Mar 22.
  PUBMED    30865830
REFERENCE   2  (bases 1 to 3831)
  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.8b00168"; journalName: "ACS Synth Biol"; date: 
            "2019-04-19- 19"; volume: "8"; issue: "4"; pages: "708-723"
COMMENT     Sequence Label: MYMH 351
FEATURES             Location/Qualifiers
     source          1..3831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    5..26
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     protein_bind    29..45
                     /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)."
     protein_bind    52..68
                     /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)."
     CDS             108..821
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(138..166)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     761..781
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      844..915
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      931..958
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    965..985
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      990..1084
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1132..1234
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             1235..1894
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1301..1320)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     oriT            2043..2151
                     /note="incP origin of transfer"
     CDS             complement(2162..2824)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      2825..3596
                     /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(3711..3797)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(3812..3831,1..3)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
ORIGIN
        1 aggagaattc gcggccgctt ctagagaatt gtgagcggat aacaattgac attgtgagcg
       61 gataacaaga tactgagcac atactagaga aagaggaggc atactagatg cgtaaaggag
      121 aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat gttaatgggc
      181 acaaattttc tgtcagtgga gagggtgaag gtgatgcaac atacggaaaa cttaccctta
      241 aatttatttg cactactgga aaactacctg ttccatggcc aacacttgtc actactttcg
      301 gttatggtgt tcaatgcttt gcgagatacc cagatcatat gaaacagcat gactttttca
      361 agagtgccat gcccgaaggt tatgtacagg aaagaactat atttttcaaa gatgacggga
      421 actacaagac acgtgctgaa gtcaagtttg aaggtgatac ccttgttaat agaatcgagt
      481 taaaaggtat tgattttaaa gaagatggaa acattcttgg acacaaattg gaatacaact
      541 ataactcaca caatgtatac atcatggcag acaaacaaaa gaatggaatc aaagttaact
      601 tcaaaattag acacaacatt gaagatggaa gcgttcaact agcagaccat tatcaacaaa
      661 atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg tccacacaat
      721 ctgccctttc gaaagatccc aacgaaaaga gagaccacat ggtccttctt gagtttgtaa
      781 cagctgctgg gattacacat ggcatggatg aactatacaa ataataatac tagagccagg
      841 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
      901 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
      961 tatatactag tagcggccgc tgcagcttgg actcctgttg atagatccag taatgacctc
     1021 agaactccat ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga
     1081 gaatccaggg gtccccaata attacgattt aaattggcga aaatgagacg ttgatcggca
     1141 cgtaagaggt tccaactttc accataatga aataagatca ctaccgggcg tattttttga
     1201 gttatcgaga ttttcaggag ctaaggaagc taaaatggag aaaaaaatca ctggatatac
     1261 caccgttgat atatcccaat ggcatcgtaa agaacatttt gaggcatttc agtcagttgc
     1321 tcaatgtacc tataaccaga ccgttcagct ggatattacg gcctttttaa agaccgtaaa
     1381 gaaaaataag cacaagtttt atccggcctt tattcacatt cttgcccgcc tgatgaatgc
     1441 tcatccggaa tttcgtatgg caatgaaaga cggtgagctg gtgatatggg atagtgttca
     1501 cccttgttac accgttttcc atgagcaaac tgaaacgttt tcatcgctct ggagtgaata
     1561 ccacgacgat ttccggcagt ttctacacat atattcgcaa gatgtggcgt gttacggtga
     1621 aaacctggcc tatttcccta aagggtttat tgagaatatg tttttcgtct cagccaatcc
     1681 ctgggtgagt ttcaccagtt ttgatttaaa cgtggccaat atggacaact tcttcgcccc
     1741 cgttttcacc atgggcaaat attatacgca aggcgacaag gtgctgatgc cgctggcgat
     1801 tcaggttcat catgccgttt gtgatggctt ccatgtcggc agaatgctta atgaattaca
     1861 acagtactgc gatgagtggc agggcggggc gtaatttgac ttttgtcctt ttccgctgca
     1921 taaccctgct tcggggtcat tatagcgatt ttttcggtat atccatcctt tttcgcacga
     1981 tatacaggat tttgccaaag ggttcgtgta gactttcctt ggtgtatcca acggcgtcag
     2041 ccgggcagga taggtgaagt aggcccaccc gcgagcgggt gttccttctt cactgtccct
     2101 tattcgcacc tggcggtgct caacgggaat cctgctctgc gaggctggcc gtaggccggc
     2161 cctaccggcg cggcagcgtt acccgtgtcg gcggctccaa cggctcgcca tcgtccagaa
     2221 aacacggctc atcgggcatc ggcaggcgct gctgcccgcg ccgttcccat tcctccgttt
     2281 cggtcaaggc tggcaggtct ggttccatgc ccggaatgcc gggctggctg ggcggctcct
     2341 cgccggggcc ggtcggtagt tgctgctcgc ccggatacag ggtcgggatg cggcgcaggt
     2401 cgccatgccc caacagcgat tcgtcctggt cgtcgtgatc aaccaccacg gcggcactga
     2461 acaccgacag gcgcaactgg tcgcggggct ggccccacgc cacgcggtca ttgaccacgt
     2521 aggccgacac ggtgccgggg ccgttgagct tcacgacgga gatccagcgc tcggccacca
     2581 agtccttgac tgcgtattgg accgtccgca aagaacgtcc gatgagcttg gaaagtgtct
     2641 tctggctgac caccacggcg ttctggtggc ccatctgcgc cacgaggtga tgcagcagca
     2701 ttgccgccgt gggtttcctc gcaataagcc cggcccacgc ctcatgcgct ttgcgttccg
     2761 tttgcaccca gtgaccgggc ttgttcttgg cttgaatgcc gatttctctg gactgcgtgg
     2821 ccatgcttat ctccatgcgg taggggtgcc gcacggttgc ggcaccatgc gcaatcagct
     2881 gcaacttttc ggcagcgcga caacaattat gcgttgcgta aaagtggcag tcaattacag
     2941 attttcttta acctacgcaa tgagctattg cggggggtgc cgcaatgagc tgttgcgtac
     3001 cccccttttt taagttgttg atttttaagt ctttcgcatt tcgccctata tctagttctt
     3061 tggtgcccaa agaagggcac ccctgcgggg ttcccccacg ccttcggcgc ggctccccct
     3121 ccggcaaaaa gtggcccctc cggggcttgt tgatcgactg cgcggccttc ggccttgccc
     3181 aaggtggcgc tgcccccttg gaacccccgc actcgccgcc gtgaggctcg gggggcaggc
     3241 gggcgggctt cgcccttcga ctgcccccac tcgcataggc ttgggtcgtt ccaggcgcgt
     3301 caaggccaag ccgctgcgcg gtcgctgcgc gagccttgac ccgccttcca cttggtgtcc
     3361 aaccggcaag cgaagcgcgc aggccgcagg ccggaggctt ttccccagag aaaattaaaa
     3421 aaattgatgg ggcaaggccg caggccgcgc agttggagcc ggtgggtatg tggtcgaagg
     3481 ctgggtagcc ggtgggcaat ccctgtggtc aagctcgtgg gcaggcgcag cctgtccatc
     3541 agcttgtcca gcagggttgt ccacgggccg agcgaagcga gccagccggt ggccgctcgc
     3601 ggccatcgtc cacatatcca cgggctggca agggagcgca gcgaccgcgc agggcgaagc
     3661 ccggagagca agcccgtagg gggggcgcgc ccagctgtct agggcggcgg atttgtccta
     3721 ctcaggagag cgttcaccga caaacaacag ataaaacgaa aggcccagtc tttcgactga
     3781 gcctttcgtt ttatttgatg cctttaatta aagcggataa caatttcaca c
//
