LOCUS       sequence-180108-        6052 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PkaiA-kaiA-ecfp expressed from NSI-SpSm vector constructed by 
            seamless assembly. Full length fusion protein expressed at WT 
            levels. ~2hr long period..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6052)
  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 6052)
  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: pAM5085
FEATURES             Location/Qualifiers
     source          1..6052
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      24..612
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     513..532
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     766..783
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    798..938
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(864..883)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(1967..2755)
                     /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             join(4451..4453,4454..5167)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4493..4514)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4754..4773)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5101..5122
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      5191..5218
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
       61 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
      121 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
      181 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
      241 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
      301 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
      361 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
      421 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
      481 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
      541 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
      601 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
      661 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
      721 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
      781 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
      841 caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
      901 acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg
      961 ctgacgcgcc ctgacgggct tgtctgctcc cgccatccgc ttacagacaa gctccgcggc
     1021 ggagcgctgc tttcttggca agcggtcgcc agccccaacg ccagggctgc cagcccgaaa
     1081 cagcggggca aggcagcttg gaagggcgat cgcagcacgg gcatggcaat gtctctctga
     1141 aggaatgcag accttattcg tacagccagg gttgaatcgt gggggtccaa tcacttagct
     1201 ctgctgggct aaaccagaga gcaatttcct gttgtgctgt ttcgattgca tccgagccat
     1261 ggatgatgtt gcggccaata ttgacaccaa aatcaccacg gatggtgccc ggttctgccg
     1321 tcagcggatt ggtagcgccg atcaacttgc gagcagccgc cacaacgcct tcgccttcca
     1381 agacgatcgc cacgatcggc ccagaggtga tgaactcgac gaggccattg aagaaggggc
     1441 gctcgcggtg gacagcatag tgctgttcgg ccagctcgcg actgggcttc agctgcttta
     1501 ggcccaccag tttgaagcct ttttgctcaa agcggccgat gatcgtaccg accaaacccc
     1561 gctgaacgcc atcgggcttg atggcaataa atgtgcgttc cacagacatc tagatagtcc
     1621 tcaagacgag gcaagcattg agcttgcctt cctatggttc gggatcactg ggattcttga
     1681 caagcgatcg cggtcacatc gctatctctt aggacttcgc agcgggcgag tcggattgac
     1741 ccggtaggga tttcgccaga tcaatgcccg tggtttgttt cagcttctcc agcaagctag
     1801 cgatttgggt agcgctgcct tccccttcgc caatcacagt gatcgactcc acgtcgatat
     1861 ctggcacggt gcctgaaagc gtgacgagca gctagcatcg cgcgcgcgcg cgcggatgct
     1921 agcggcgcgc caaaaaaaag cccgctcatt aggcgggctg gccggccttg ccgactacct
     1981 tggtgatctc gcctttcacg tagtggacaa attcttccaa ctgatctgcg cgcgaggcca
     2041 agcgatcttc ttcttgtcca agataagcct gtctagcttc aagtatgacg ggctgatact
     2101 gggccggcag gcgctccatt gcccagtcgg cagcgacatc cttcggcgcg attttgccgg
     2161 ttactgcgct gtaccaaatg cgggacaacg taagcactac atttcgctca tcgccagccc
     2221 agtcgggcgg cgagttccat agcgttaagg tttcatttag cgcctcaaat agatcctgtt
     2281 caggaaccgg atcaaagagt tcctccgccg ctggacctac caaggcaacg ctatgttctc
     2341 ttgcttttgt cagcaagata gccagatcaa tgtcgatcgt ggctggctcg aagatacctg
     2401 caagaatgtc attgcgctgc cattctccaa attgcagttc gcgcttagct ggataacgcc
     2461 acggaatgat gtcgtcgtgc acaacaatgg tgacttctac agcgcggaga atctcgctct
     2521 ctccagggga agccgaagtt tccaaaaggt cgttgatcaa agctcgccgc gttgtttcat
     2581 caagccttac ggtcaccgta accagcaaat caatatcact gtgtggcttc aggccgccat
     2641 ccactgcgga gccgtacaaa tgtacggcca gcaacgtcgg ttcgagatgg cgctcgatga
     2701 cgccaactac ctctgatagt tgagtcgata cttcggcgat caccgcttcc ctcatgatgt
     2761 ttaactttgt tttagggcga ctgccctgct gcgtaacatc gttgctgctc cataacatca
     2821 aacatcgacc cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata gactgtaccc
     2881 caaaaaaaca gtcataacaa gccatgaaaa ccgccactgc gccgttacca ccgctgcgtt
     2941 cggtcaaggt tctggaccag ttgcgtgagc gcatacgcta cttgcattac agcttacgaa
     3001 ccgaacaggc ttatgtccac tgggttcgtg ccttcatccg tttccacggt gtgcgtcacc
     3061 cggcaacctt gggcagcagc gaagtcgagg catttctgtc ctggctggcg aacgagcgca
     3121 aggtttcggt ctccacgcat tcgaaaccgg tatcccgccg ccgccgccgg ataccggtta
     3181 atagtacctg tgaagtgaaa aatggcgcac attgtgcgac atttttttct cgagcaggta
     3241 gtcggccaat aacccaggga ttttccaagt ctgcgtcgag gcgatcgaga tcgtagaacc
     3301 gaccgggttc aacccagagt tgcttaccgc ttgcttcgat gattgcgtaa gccatgcatt
     3361 ttcagccgta gaggtggcta cacctgccga gggcagagag agcacttgaa gaaacctgtt
     3421 ccgagccgcc ttttaaacaa cagccctcta tcatctcaga aacttttgag aactgtcaaa
     3481 ctgtggacaa agcgatcgcg gcctcatgag ctgcagtgct aggctaaatt aaatttttcc
     3541 tttgtccaga gattaatctg tctgcagact cagtcctgac aggagcgact gcgtgctctc
     3601 gcaaattgca atctgcattt gggtggaatc gacggcaatt ttgcaggatt gccagcgggc
     3661 gctgtcggcc gatcgctatc aactccaagt ctgtgagtct ggcgaaatgc tcttggagta
     3721 tgcccaaacc catcgtgacc aaatcgactg cctgatttta gtggcagcca atcccagctt
     3781 cagggcagtt gttcagcagc tctgctttga gggagtggtg gtaccagcga ttgtcgtagg
     3841 cgatcgcgac agtgaggatc ccgatgaacc agccaaagaa cagctctatc acagcgctga
     3901 actgcacctc ggtatccatc agctcgagca attgccctac caagttgatg ctgcactggc
     3961 tgaatttctg cgcttagccc cggtcgagac catggccgac cacatcatgc tgatgggggc
     4021 caaccacgat cccgagctat cgagccagca gcgggacctc gctcagcgac tacaagagcg
     4081 cctaggctat ctcggggtct actacaagcg tgatcccgat cgctttctgc gcaacctacc
     4141 cgcctacgaa agccaaaagc tgcaccaagc gatgcagact agctatcgtg aaatcgtttt
     4201 gagctatttt tcgccgaata gcaacctcaa ccagagcatt gacaacttcg tcaacatggc
     4261 tttctttgcc gatgttccag tcaccaaagt ggtagaaatt cacatggagc tgatggacga
     4321 gtttgccaag aagctccgcg tagagggacg ttcagaggac attttgctgg attatcggct
     4381 gactttaatt gatgtaattg cacatctttg tgagatgtat cgacggtcta tcccacgaga
     4441 aaccgcggca gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     4501 gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc
     4561 cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg
     4621 gcccaccctc gtgaccaccc tgacctgggg cgtgcagtgc ttcagccgct accccgacca
     4681 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     4741 catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga
     4801 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     4861 ggggcacaag ctggagtaca actacatcag ccacaacgtc tatatcaccg ccgacaagca
     4921 gaagaacggc atcaaggcca acttcaagat ccgccacaac atcgaggacg gcagcgtgca
     4981 gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga
     5041 caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca
     5101 catggtcctg ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta
     5161 caagtaaaaa tgaaggagct ccccggcagg agaaggccat cctgacggat ggcctttttg
     5221 aagctttcta gaatcggggg cccccggggg gattctagat tctggagcag gaagatgtcg
     5281 cgggcattag caccagcggt ctgccaagcc tccgccagcc gttgggtccc ttccgcttga
     5341 gcttttccat cttcgacgat acgggcggcg gccccccgcg cttccgcgat cgcccgttta
     5401 caagctgcct cagctggggc gatcacatcg gcttgaagtt gctgctgcac ctgtttgatc
     5461 cgctcctgct gcacagggag ttctgcttgg ctacgagcga cttcggtagc aatgtccgct
     5521 tcagcttcgg ccaccaccgc ttcgcgccgc gtcaacgcat cctgaatccg gcgctcggcc
     5581 tcggcttggg cgatcgctac atcgcgatcg atccgacgca gggccgtgat cttgtcattt
     5641 tcggccgttt ggatcgcaga ggcagcctgg gcatcggctt cagcaattcg ggcatctcgc
     5701 tgcagatcag cccgctgctt gcgtccacta gccgagagat aaccgacctc atcggaaatg
     5761 ttctggactt gcagcgtatc gaggactaga cccagctgct caaggtcatc ctccgcctct
     5821 tccagcagac ttttggcaaa ggcaattttg tcctcgttga tctgctccgg cgtgaggctg
     5881 gctaaaacac cacgcaagtt gccttcgagg gtctccttgg caatttgctc gatttcctta
     5941 cggtttttgc caagcagccg ctcgatcgcg ttgtggatgg tcggttcttc cccagcaatc
     6001 ttgatattgg caacgccttc aacagtcagg ggaatgccga gcgtcagacc cc
//
