LOCUS       sequence-180104-        7793 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ptrc-yfp-kaiC expressed from NSI-SpSm vector constructed by seamless
            assembly. Subcloned from pAM5080. Functional fusion expressed as 
            full-length fusion protein expressed at WT levels..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7793)
  AUTHORS   Cohen SE, Erb ML, Selimkhanov J, Dong G, Hasty J, Pogliano J, Golden
            SS
  TITLE     Dynamic localization of the cyanobacterial circadian clock proteins.
  JOURNAL   Curr Biol. 2014 Aug 18;24(16):1836-44. doi: 
            10.1016/j.cub.2014.07.036. Epub 2014 Aug 7.
  PUBMED    25127213
REFERENCE   2  (bases 1 to 7793)
  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: 
            "10.1016/j.cub.2014.07.036"; journalName: "Curr Biol"; date: 
            "2014-08-18- 18"; volume: "24"; issue: "16"; pages: "1836-44"
COMMENT     Sequence Label: pAM5081
FEATURES             Location/Qualifiers
     source          1..7793
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    775..915
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(841..860)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(1944..2732)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     CDS             complement(3241..4323)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4285..4304
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4324..4401)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     promoter        join(4469..4474,4475..4491,4492..4498)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    4506..4522
                     /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             join(4558..4560,4561..4563,4564..5274)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVRFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4603..4624)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4864..4883)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5211..5232
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             5275..5304
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      6909..6936
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
      781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt gcactctcag
      841 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
      901 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
      961 ctgctcccgc catccgctta cagacaagct ccgcggcgga gcgctgcttt cttggcaagc
     1021 ggtcgccagc cccaacgcca gggctgccag cccgaaacag cggggcaagg cagcttggaa
     1081 gggcgatcgc agcacgggca tggcaatgtc tctctgaagg aatgcagacc ttattcgtac
     1141 agccagggtt gaatcgtggg ggtccaatca cttagctctg ctgggctaaa ccagagagca
     1201 atttcctgtt gtgctgtttc gattgcatcc gagccatgga tgatgttgcg gccaatattg
     1261 acaccaaaat caccacggat ggtgcccggt tctgccgtca gcggattggt agcgccgatc
     1321 aacttgcgag cagccgccac aacgccttcg ccttccaaga cgatcgccac gatcggccca
     1381 gaggtgatga actcgacgag gccattgaag aaggggcgct cgcggtggac agcatagtgc
     1441 tgttcggcca gctcgcgact gggcttcagc tgctttaggc ccaccagttt gaagcctttt
     1501 tgctcaaagc ggccgatgat cgtaccgacc aaaccccgct gaacgccatc gggcttgatg
     1561 gcaataaatg tgcgttccac agacatctag atagtcctca agacgaggca agcattgagc
     1621 ttgccttcct atggttcggg atcactggga ttcttgacaa gcgatcgcgg tcacatcgct
     1681 atctcttagg acttcgcagc gggcgagtcg gattgacccg gtagggattt cgccagatca
     1741 atgcccgtgg tttgtttcag cttctccagc aagctagcga tttgggtagc gctgccttcc
     1801 ccttcgccaa tcacagtgat cgactccacg tcgatatctg gcacggtgcc tgaaagcgtg
     1861 acgagcagct agcatcgcgc gcgcgcgcgc ggatgctagc ggcgcgccaa aaaaaagccc
     1921 gctcattagg cgggctggcc ggccttgccg actaccttgg tgatctcgcc tttcacgtag
     1981 tggacaaatt cttccaactg atctgcgcgc gaggccaagc gatcttcttc ttgtccaaga
     2041 taagcctgtc tagcttcaag tatgacgggc tgatactggg ccggcaggcg ctccattgcc
     2101 cagtcggcag cgacatcctt cggcgcgatt ttgccggtta ctgcgctgta ccaaatgcgg
     2161 gacaacgtaa gcactacatt tcgctcatcg ccagcccagt cgggcggcga gttccatagc
     2221 gttaaggttt catttagcgc ctcaaataga tcctgttcag gaaccggatc aaagagttcc
     2281 tccgccgctg gacctaccaa ggcaacgcta tgttctcttg cttttgtcag caagatagcc
     2341 agatcaatgt cgatcgtggc tggctcgaag atacctgcaa gaatgtcatt gcgctgccat
     2401 tctccaaatt gcagttcgcg cttagctgga taacgccacg gaatgatgtc gtcgtgcaca
     2461 acaatggtga cttctacagc gcggagaatc tcgctctctc caggggaagc cgaagtttcc
     2521 aaaaggtcgt tgatcaaagc tcgccgcgtt gtttcatcaa gccttacggt caccgtaacc
     2581 agcaaatcaa tatcactgtg tggcttcagg ccgccatcca ctgcggagcc gtacaaatgt
     2641 acggccagca acgtcggttc gagatggcgc tcgatgacgc caactacctc tgatagttga
     2701 gtcgatactt cggcgatcac cgcttccctc atgatgttta actttgtttt agggcgactg
     2761 ccctgctgcg taacatcgtt gctgctccat aacatcaaac atcgacccac ggcgtaacgc
     2821 gcttgctgct tggatgcccg aggcatagac tgtaccccaa aaaaacagtc ataacaagcc
     2881 atgaaaaccg ccactgcgcc gttaccaccg ctgcgttcgg tcaaggttct ggaccagttg
     2941 cgtgagcgca tacgctactt gcattacagc ttacgaaccg aacaggctta tgtccactgg
     3001 gttcgtgcct tcatccgttt ccacggtgtg cgtcacccgg caaccttggg cagcagcgaa
     3061 gtcgaggcat ttctgtcctg gctggcgaac gagcgcaagg tttcggtctc cacgcattcg
     3121 aaaccggtat cccgccgccg ccgccggata ccggttaata gtacctgtga agtgaaaaat
     3181 ggcgcacatt gtgcgacatt tttttctcga gcaggtagtc ggccaataac ccagggattt
     3241 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     3301 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
     3361 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
     3421 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
     3481 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
     3541 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
     3601 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
     3661 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
     3721 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
     3781 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
     3841 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
     3901 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
     3961 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
     4021 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
     4081 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
     4141 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
     4201 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     4261 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     4321 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     4381 tttgcaccat tcgatggtgt caacgtaaat gcatgccgct tcgccttcgc gcgcgaattg
     4441 atctgcggcg gccgcctgaa atgagctgtt gacaattaat catccggctc gtataatgtg
     4501 tggaattgtg agcggataac aatttcacac gcatgcggat cctaaggagg aaaaaaaatg
     4561 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     4621 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     4681 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     4741 gtgaccacct tcggctacgg cctgcagtgc ttcgcccgct accccgacca catgaagcag
     4801 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     4861 aaggacgacg gcaactacaa gacccgcgcc gaggtgaggt tcgagggcga caccctggtg
     4921 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     4981 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     5041 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     5101 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     5161 ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     5221 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caaggaacag
     5281 aagcttatca gcgaagaaga tctgggctct ggatcgggtt caggaatgac ttccgctgag
     5341 atgactagcc ctaataataa ttctgagcac caagccatcg ctaagatgcg cacgatgatt
     5401 gaaggctttg atgatattag tcatggcggt cttccaatcg ggcgatcgac cctcgttagt
     5461 ggtacttcag gaaccggcaa gacccttttt tctattcaat ttctctataa cggtattatc
     5521 gagtttgatg agcctggggt tttcgttact ttcgaagaga ccccgcaaga tatcattaaa
     5581 aacgcccgta gttttggctg ggatttagcc aagctggtcg atgagggcaa actatttatt
     5641 cttgatgctt cacccgatcc agaaggtcaa gaggttgttg gcggcttcga tctctctgct
     5701 ctgattgagc ggattaatta tgcaattcaa aagtatcgag cgcggcgggt ttcaattgac
     5761 tcggtcacgt ccgttttcca gcaatatgat gcctcttctg tggttcgccg cgaactcttt
     5821 cggttggtag ctcgcctaaa acaaattggg gcaactacgg tcatgaccac cgagcgtatc
     5881 gaggaatatg gcccgatcgc tcgttacggt gttgaggaat ttgtctccga taacgtcgtg
     5941 attctccgca acgttttgga aggggagcgc cgtcgccgca ccctcgaaat cctcaagcta
     6001 cgtggcacca gccacatgaa aggggaatat ccgttcacga ttacggatca tggcatcaat
     6061 atcttcccgc tcggggcaat gcgccttacg cagcgatcgt cgaacgtgcg tgtttcatct
     6121 ggtgtcgtcc gactcgatga aatgtgtggt gggggcttct ttaaggactc aatcattctg
     6181 gcaactggcg ctacaggcac tggtaaaact ctgttagtta gccgtttcgt tgagaatgct
     6241 tgtgctaaca aagagcgggc gattctgttc gcttatgaag agtcacgagc tcagctgctc
     6301 cgcaatgcct attcatgggg aatggacttt gaggagatgg agcgccaaaa cctcctcaaa
     6361 attgtttgcg cctatcctga atctgcaggt cttgaagacc atttgcagat tattaagtcg
     6421 gagatcaatg actttaagcc agctcgtatt gcaatcgact ccctctctgc tttggcgcgg
     6481 ggcgttagca acaatgcctt ccgccaattt gtaattggtg tcactggcta cgcgaaacaa
     6541 gaagaaatca cgggactatt cacaaatacc agtgatcaat ttatgggagc gcattcgatt
     6601 actgactccc atatctcaac aattacggat acgattatct tgctccaata cgtcgagatt
     6661 cgtggcgaaa tgtcccgcgc cattaacgtc ttcaagatgc gcggatcttg gcatgacaaa
     6721 gcaatccgcg aattcatgat cagcgacaaa gggccggaca tcaaggattc tttccggaac
     6781 tttgagcgga ttatttcagg ttcgccaaca cggattaccg tcgatgagaa aagcgaactc
     6841 tcgcgaattg tgcgcggcgt tcaagaaaaa gggccggaga gctagaaatg aaggagctcc
     6901 ccggcaggag aaggccatcc tgacggatgg cctttttgaa gctttctaga atcgggggcc
     6961 cccgggggga ttctagattc tggagcagga agatgtcgcg ggcattagca ccagcggtct
     7021 gccaagcctc cgccagccgt tgggtccctt ccgcttgagc ttttccatct tcgacgatac
     7081 gggcggcggc cccccgcgct tccgcgatcg cccgtttaca agctgcctca gctggggcga
     7141 tcacatcggc ttgaagttgc tgctgcacct gtttgatccg ctcctgctgc acagggagtt
     7201 ctgcttggct acgagcgact tcggtagcaa tgtccgcttc agcttcggcc accaccgctt
     7261 cgcgccgcgt caacgcatcc tgaatccggc gctcggcctc ggcttgggcg atcgctacat
     7321 cgcgatcgat ccgacgcagg gccgtgatct tgtcattttc ggccgtttgg atcgcagagg
     7381 cagcctgggc atcggcttca gcaattcggg catctcgctg cagatcagcc cgctgcttgc
     7441 gtccactagc cgagagataa ccgacctcat cggaaatgtt ctggacttgc agcgtatcga
     7501 ggactagacc cagctgctca aggtcatcct ccgcctcttc cagcagactt ttggcaaagg
     7561 caattttgtc ctcgttgatc tgctccggcg tgaggctggc taaaacacca cgcaagttgc
     7621 cttcgagggt ctccttggca atttgctcga tttccttacg gtttttgcca agcagccgct
     7681 cgatcgcgtt gtggatggtc ggttcttccc cagcaatctt gatattggca acgccttcaa
     7741 cagtcagggg aatgccgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
