LOCUS       sequence-151985-        5859 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive CcaS and CcaR with gfpmut3 under the cpcG2-Delta-59 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5859)
  AUTHORS   Milias-Argeitis A, Rullan M, Aoki SK, Buchmann P, Khammash M
  TITLE     Automated optogenetic feedback control for precise and robust 
            regulation of gene expression and cell growth.
  JOURNAL   Nat Commun. 2016 Aug 26;7:12546. doi: 10.1038/ncomms12546.
  PUBMED    27562138
REFERENCE   2  (bases 1 to 5859)
  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: "2016-08-26"; volume: "7:12546. doi"; pages: " 
            10.1038/ncomms12546"
COMMENT     Sequence Label: pSKA413
FEATURES             Location/Qualifiers
     source          1..5859
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(1..48)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(52..75)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(2365..2388)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3298..4011
                     /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(3328..3356)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3951..3971
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      4026..4112
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      complement(4277..4865)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4357..4376)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4953..5047)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(5068..5727)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5642..5661
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(5728..5830)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
ORIGIN
        1 caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctagtt acttgtcatc
       61 gtcgtccttg tagtcagctc gaggcaaatg gttatagcgt tttaaacaga tagtaactgt
      121 tgttccctgg tcaactgcac tttttagcaa gatactgcca ctgtgtaagt caacacattt
      181 cttggcaacc attaaaccga gtcctgttcc cgtaatattt ctgacatttt tgccccgatg
      241 aaagggttca aaaatttgct tttggtcctc tggcgaaatg ccaatgccct ggtcggtgac
      301 ttcaaaaata atctgttccg aatctaggct tagggcaatt ttaatctgcc ctcccccggg
      361 agagtattta atcgcattag ataacagatt agataaaata gaccgcacta acctttcatc
      421 caccaatgcc ttcgtatctt gagcgctaca aataaagtca aaataatatt ggtcactgac
      481 acttaattga atttcttcga taaattgctg gaacaataat ttcaaatcta accaattagg
      541 attaaattcc aatttgcccg cttcggcacg gttaatgatt aaaatatcat ccaggagctg
      601 taccatattt ttcacggaat tttgaatacg gtgtaagttc cggctacgct tatcgggatc
      661 aagccaggcc acttcagaat tttccagtaa ttgggcagca gctaaggccg tactgagggg
      721 agtacgaaat tcatgggaag ccatggagaa aaaacgagtt tttaggcggc ttaattcttt
      781 ttcccgttct aaagctaggc ggacttcctc cgtgcgtcgg cgctcagtga catccctttg
      841 cacagcaatc caatgggtgt aaaaacctgt tttattggca acgggcacca gactaaattc
      901 cacccaaaac tcactgccat ccttacgata attaatcact tcaacggtaa ctgattgcca
      961 ttgactaatg gcttgccgca ccctatctaa ttcagtgcga ctagtttttg gtccctgtaa
     1021 aactcggggg gttttgccta gcatttcttc agcagtataa ccggtgattt tagtaaatgc
     1081 ttcattgaca tagagaattc tcggcccggg atcatcaatg ggctccgctt ccgtaatcac
     1141 caccgcatca ttagtattaa ccacaacgga ttgtagcaaa cgaatttgtt ctgcctgccg
     1201 cttttgctcg gtaatatcgg cgacaaccgc caccaatccg gcaatatttt cttcactatc
     1261 caatagggga gcagcggaaa ggacaatttc aatccaacta ccatcttttt tttgacattt
     1321 taattctagg ctaggggaaa ccatgcctga taaaactttc tgtttaaatt gctgataatc
     1381 ttccagcaaa atgttggatg ttaataattc tggatgggca ataatttctg tttctgtcca
     1441 accaaaaata cgttctgctg taggattcca aatctgaatt tggtctgcta aattaagggt
     1501 aaaaataccc ctgggggagg ccgcaatcag ggattgcaga gtatggttag tgtggcggag
     1561 agcggcttcc gttttttgtc gctcactgat ttccattctt agggattgat tggtggcggc
     1621 aagttgctgg gtgcgttttt cgactcggtt ttccaaatct ttattgagtt gctgtaattg
     1681 ctcatataat tccgattgtt ggatggcgat cgcgacctga ttggctagct gtttcattaa
     1741 ctccacttcc cacggttgcc atggccgggt aaaagcacat tgatgggtaa ttaacagccc
     1801 ccaaagatag ggaaattctg attcattatc tagggaagaa gcacgattat gttgaagaat
     1861 gggcactact aattttgatt tcacgccaaa ttgtttgacg aaatcagcta ggcaaatctc
     1921 tatgtcatcc tggtcaatgt cattaatggc ccgtacttta cctttggtgt aggcttgatg
     1981 gtattcaacg ggaaaaactt catcggaaaa ggtccgccct aaaatactag gataattggc
     2041 attcaccgat tccgtaatgg cgctgcccgt gccatcttgc caaatgcgat aaattagaac
     2101 tcgatcaact tgcaacaggg ttttaacttc agcgacggtg gtatttaaaa ccgtttccaa
     2161 gtttaaagat tggcggatat gttgggtaat ttggttaata aatcgttccc gttgggtttg
     2221 ttgcttgatg ctaatttcaa tcctgcggcg ttcttggttt tgtctgtgcc ataaataaca
     2281 ggtcatggcg atcgccagaa aaacaaattc gattggaatt agaaatttgc ccatatatta
     2341 gcgaatccca gtagaaaaac cctacttgtc atcgtcgtcc ttgtagtcgt ttttcccttg
     2401 gcacaaagat ttttccgtta aattggccag acgataccca atgccatgga ccgtttcaat
     2461 ggcatctgct gataaaccgg cactttttaa tttttgtcgc aaactccgca catgcacctt
     2521 aaccgtatct tcctctgggg gactctccaa cttccagata ctatcgataa tcatgctccg
     2581 acttagcacc cgacggccat tgcggagtag taattccaga atgctgtatt ccttgcgggt
     2641 caaagacaaa acctcattgt cataactaac ttcataggtg cttggatcca acctgattgg
     2701 cccccactct aaaactggtt ggcacgttgc acaaccccga cgcaacaaag ctcgcaccct
     2761 ggcaaataac tcccccaaat ccactggctt gaccacataa tcatccgccc ccgcatccaa
     2821 gcccgtgatt ttatcattga tcgtatccct ggctgtcatc atcaaaattg gcattaaata
     2881 actgtgcgat cgccattttt gacagagggt aatcccgtcc aactccggca gcatcacatc
     2941 caaaataacg aggtcatatt ccagtcggga ggcatagtcc caggcgaggg aagcgtcggt
     3001 ggcaatatca acggtgtaaa gctggtcact caatgcttca gcaagggttt ccgccagcgg
     3061 caaatcatcc tccactaaaa gaattctcat agcccatcct gcttttctct atcaacctca
     3121 gcttacctga aggggtgaac aggtctgggt taattcatgt tgcgaaatgt aacagtttta
     3181 gtcgcatcag ctaactttcc gatttcttta cgattttctc ccccttttct tcaattttac
     3241 tttgttagga tcgcattcta actttatcta agggacaggc acgtaaggca ccatataatg
     3301 cgtaaaggag aagaactttt cactggagtt gtcccaattc ttgttgaatt agatggtgat
     3361 gttaatgggc acaaattttc tgtcagtgga gagggtgaag gtgatgcaac atacggaaaa
     3421 cttaccctta aatttatttg cactactgga aaactacctg ttccatggcc aacacttgtc
     3481 actactttcg gttatggtgt tcaatgcttt gcgagatacc cagatcatat gaaacagcat
     3541 gactttttca agagtgccat gcccgaaggt tatgtacagg aaagaactat atttttcaaa
     3601 gatgacggga actacaagac acgtgctgaa gtcaagtttg aaggtgatac ccttgttaat
     3661 agaatcgagt taaaaggtat tgattttaaa gaagatggaa acattcttgg acacaaattg
     3721 gaatacaact ataactcaca caatgtatac atcatggcag acaaacaaaa gaatggaatc
     3781 aaagttaact tcaaaattag acacaacatt gaagatggaa gcgttcaact agcagaccat
     3841 tatcaacaaa atactccaat tggcgatggc cctgtccttt taccagacaa ccattacctg
     3901 tccacacaat ctgccctttc gaaagatccc aacgaaaaga gagaccacat ggtccttctt
     3961 gagtttgtaa cagctgctgg gattacacat ggcatggatg aactatacaa ataatctaga
     4021 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     4081 tgtcggtgaa cgctctcctg agtaggacaa atccgccgcc ctagacctag ggcgttcggc
     4141 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4201 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4261 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4321 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4381 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4441 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4501 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4561 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4621 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4681 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     4741 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4801 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4861 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4921 gttaagggat tttggtcatg actagtgctt ggattctcac caataaaaaa cgcccggcgg
     4981 caaccgagcg ttctgaacaa atccagatgg agttctgagg tcattactgg atctatcaac
     5041 aggagtccaa gcgagctcga tatcaaatta cgccccgccc tgccactcat cgcagtactg
     5101 ttgtaattca ttaagcattc tgccgacatg gaagccatca caaacggcat gatgaacctg
     5161 aatcgccagc ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac
     5221 gggggcgaag aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca
     5281 gggattggct gagacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt
     5341 ttcaccgtaa cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg
     5401 gtattcactc cagagcgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg
     5461 gtgaacacta tcccatatca ccagctcacc gtctttcatt gccatacgaa attccggatg
     5521 agcattcatc aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt
     5581 ctttacggtc tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg
     5641 agcaactgac tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt
     5701 ggtatatcca gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgataa
     5761 ctcaaaaaat acgcccggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac
     5821 gtgccgatca acgtctcatt ttcgccagat atcgacgtc
//
