LOCUS       sequence-189868-        7322 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Ptrc-yfp-kaiC expressed from NSII-Km vector constructed by seamless 
            assembly. LacI expressed in opposite direction as Ptrc promoter. 
            Functional fusion expres.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7322)
  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 7322)
  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: pAM5080
FEATURES             Location/Qualifiers
     source          1..7322
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      504..1092
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     993..1012
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1246..1263
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    1278..1418
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1344..1363)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(2552..3367)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2552..3367)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3275..3294
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3359..3378
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             complement(3592..4674)
                     /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     4636..4655
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4675..4752)
                     /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(4820..4825,4826..4842,4843..4849)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    4857..4873
                     /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(4909..4911,4912..4914,4915..5625)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVRFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4954..4975)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5215..5234)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5562..5583
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             5626..5655
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      7260..7287
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 tctagagttg ctaccaagta gtgacccgct tcgtgatgca aaatccgctg acgatattcg
       61 ggcgatcgct gctgaatgcc atcgagcagt aacgtggcac cccgcccctg ccaagtcacc
      121 gcatccagac tgaacagcac caagaggcta aaacccaatc ccgccggtag cagcggagaa
      181 ctacccagca ttggtcccac caaagctaat gccgtcgtgg taaaaatcgc gatcgccgtc
      241 agactcaagc ccagttcgct catgcttcct catctaggtc acagtcttcg gcgatcgcat
      301 cgatctgatg ctgcagcaag cgttttccat accggcgatc gcgccgtcgc cctttcgctg
      361 ccgtggcccg cttacgagct cgtttatcga ccacgatcgc atccaaatcc gcgatcgctt
      421 cccagtccgg caattcagtc tggggcgtcc gtttcattaa tcctgatagc gtcagacccc
      481 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
      541 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
      601 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
      661 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
      721 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
      781 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
      841 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
      901 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
      961 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     1021 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     1081 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     1141 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     1201 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     1261 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     1321 caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     1381 acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg ccaacacccg
     1441 ctgacgcgcc ctgacgggct tgtctgctcc cgccatccgc ttacagacaa gctccgcggg
     1501 ccacgatttg agggacgaat cttcagcgtg actgaagtac ctcgcggcaa gccatttccc
     1561 gatgtctttt tgttggccgc cgatcgcttc ggggttaatc ctacggcctg cgctgtgatc
     1621 gaagacaccc ccttgggagt agcggcaggc gtggcggcag gaatgcaagt gtttggctac
     1681 gcgggttcca tgcccgcttg gcgtctgcaa gaagccggtg cccatctcat ttttgacgat
     1741 atgcgactgc tgcccagtct gctccaatcg tcgccaaaag ataactccac agcattgccc
     1801 aatccctaac ccctgctcgc gccgcaacta cacactaaac cgttcctgcg cgatcgctct
     1861 tactgttgat ggctcgtgct taaaaacaat gcaaccctaa ccgtttcagc tggtgatttt
     1921 cggacgattt ggcttacagg gataactgag agtcaacagc ctctgtccgt cattgcacac
     1981 ccatccatgc actggggact tgactcatgc tgaatcacat ttcccttgtc cattgggcga
     2041 gaggggaggg gaatcttctg gactcttcac taagcggcga tcgcaggttc ttctacccaa
     2101 gcagtggcga tcgcttgatt gcagtcttca atgctggcct ctgcagccat cgccgccacc
     2161 aaagcatcgt aggcgggacg ttgttgctcc agtaaagtct tcgcccgtaa caatccccag
     2221 cgactgcgta aatccgcttc ggcaggattg cgatcgagtt gccgccacag ttgtttccac
     2281 tgggcgcgat cgtcagctcc cccttccacg ttgccgtaga ccagttgctc tgccgctgca
     2341 ccggccatca acacctgaca ccactgttcc agcgatcgct gactgagttg cccctgtgcg
     2401 gcttcggctt ctagcgcagc tgcttggaac tgcacacccc cgctagcatc gcgcgcgcgc
     2461 gcgcggatgc tagccaatgg cttattttcc tgctaattgg taataccatg gtaccgctct
     2521 gccagtgtta caaccaatta accaattctg attagaaaaa ctcatcgagc atcaaatgaa
     2581 actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta
     2641 atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg
     2701 cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt
     2761 tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagcttat
     2821 gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg
     2881 catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc
     2941 tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg
     3001 catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc
     3061 cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg
     3121 tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat
     3181 tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca
     3241 atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata
     3301 aatcagcatc catgttggaa tttaatcgcg gcctggagca agacgtttcc cgttgaatat
     3361 ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgatg
     3421 atatattttt atcttgtgca atgtaacatc agagattttg agacacaacg tggaccggta
     3481 tcccgccgcc gccgccggat accggttaat agtacctgtg aagtgaaaaa tggcgcacat
     3541 tgtgcgacat ttttttctcg agcaggtagt cggccaataa cccagggatt ttcactgccc
     3601 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     3661 agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa
     3721 cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt
     3781 ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct
     3841 gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc
     3901 ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg
     3961 aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc
     4021 gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc
     4081 cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg
     4141 acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat
     4201 actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc
     4261 agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac
     4321 gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac
     4381 catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat
     4441 ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt
     4501 gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc
     4561 cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt
     4621 ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac
     4681 caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcacca
     4741 ttcgatggtg tcaacgtaaa tgcatgccgc ttcgccttcg cgcgcgaatt gatctgcggc
     4801 ggccgcctga aatgagctgt tgacaattaa tcatccggct cgtataatgt gtggaattgt
     4861 gagcggataa caatttcaca cgcatgcgga tcctaaggag gaaaaaaaat ggtgagcaag
     4921 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     4981 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     5041 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     5101 ttcggctacg gcctgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
     5161 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     5221 ggcaactaca agacccgcgc cgaggtgagg ttcgagggcg acaccctggt gaaccgcatc
     5281 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     5341 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     5401 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     5461 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagctac
     5521 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     5581 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaaggaaca gaagcttatc
     5641 agcgaagaag atctgggctc tggatcgggt tcaggaatga cttccgctga gatgactagc
     5701 cctaataata attctgagca ccaagccatc gctaagatgc gcacgatgat tgaaggcttt
     5761 gatgatatta gtcatggcgg tcttccaatc gggcgatcga ccctcgttag tggtacttca
     5821 ggaaccggca agaccctttt ttctattcaa tttctctata acggtattat cgagtttgat
     5881 gagcctgggg ttttcgttac tttcgaagag accccgcaag atatcattaa aaacgcccgt
     5941 agttttggct gggatttagc caagctggtc gatgagggca aactatttat tcttgatgct
     6001 tcacccgatc cagaaggtca agaggttgtt ggcggcttcg atctctctgc tctgattgag
     6061 cggattaatt atgcaattca aaagtatcga gcgcggcggg tttcaattga ctcggtcacg
     6121 tccgttttcc agcaatatga tgcctcttct gtggttcgcc gcgaactctt tcggttggta
     6181 gctcgcctaa aacaaattgg ggcaactacg gtcatgacca ccgagcgtat cgaggaatat
     6241 ggcccgatcg ctcgttacgg tgttgaggaa tttgtctccg ataacgtcgt gattctccgc
     6301 aacgttttgg aaggggagcg ccgtcgccgc accctcgaaa tcctcaagct acgtggcacc
     6361 agccacatga aaggggaata tccgttcacg attacggatc atggcatcaa tatcttcccg
     6421 ctcggggcaa tgcgccttac gcagcgatcg tcgaacgtgc gtgtttcatc tggtgtcgtc
     6481 cgactcgatg aaatgtgtgg tgggggcttc tttaaggact caatcattct ggcaactggc
     6541 gctacaggca ctggtaaaac tctgttagtt agccgtttcg ttgagaatgc ttgtgctaac
     6601 aaagagcggg cgattctgtt cgcttatgaa gagtcacgag ctcagctgct ccgcaatgcc
     6661 tattcatggg gaatggactt tgaggagatg gagcgccaaa acctcctcaa aattgtttgc
     6721 gcctatcctg aatctgcagg tcttgaagac catttgcaga ttattaagtc ggagatcaat
     6781 gactttaagc cagctcgtat tgcaatcgac tccctctctg ctttggcgcg gggcgttagc
     6841 aacaatgcct tccgccaatt tgtaattggt gtcactggct acgcgaaaca agaagaaatc
     6901 acgggactat tcacaaatac cagtgatcaa tttatgggag cgcattcgat tactgactcc
     6961 catatctcaa caattacgga tacgattatc ttgctccaat acgtcgagat tcgtggcgaa
     7021 atgtcccgcg ccattaacgt cttcaagatg cgcggatctt ggcatgacaa agcaatccgc
     7081 gaattcatga tcagcgacaa agggccggac atcaaggatt ctttccggaa ctttgagcgg
     7141 attatttcag gttcgccaac acggattacc gtcgatgaga aaagcgaact ctcgcgaatt
     7201 gtgcgcggcg ttcaagaaaa agggccggag agctagaaat gaaggagctc cccggcagga
     7261 gaaggccatc ctgacggatg gcctttttga agctttctag aatcgggggc ccccgggggg
     7321 at
//
