LOCUS       sequence-435614-        8399 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For creating CyanoTag vectors by Golden Gate cloning (BspQI 
            version).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8399)
  TITLE     CyanoTag plasmids, for native C-terminal fluorescent tagging of 
            proteins in cyanobacteria (S. elongatus 7942)
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8399)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pLM433
FEATURES             Location/Qualifiers
     source          1..8399
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             718..783
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     terminator      791..837
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      876..903
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     primer_bind     complement(1042..1061)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1053..1868
                     /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             1053..1868
                     /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     complement(1126..1145)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(2881..4164)
                     /codon_start=1
                     /gene="codA"
                     /product="E. coli cytosine deaminase"
                     /label=codA
                     /note="confers sensitivity to 5-fluorocytosine"
                     /translation="MSNNALQTIINARLPGEEGLWQIHLQDGKISAIDAQSGVMPITEN
                     SLDAEQGLVIPPFVEPHIHLDTTQTAGQPNWNQSGTLFEGIERWAERKALLTHDDVKQR
                     AWQTLKWQIANGIQHVRTHVDVSDATLTALKAMLEVKQEVAPWIDLQIVAFPQEGILSY
                     PNGEALLEEALRLGADVVGAIPHFEFTREYGVESLHKTFALAQKYDRLIDVHCDEIDDE
                     QSRFVETVAALAHHEGMGARVTASHTTAMHSYNGAYTSRLFRLLKMSGINFVANPLVNI
                     HLQGRFDTYPKRRGITRVKEMLESGINVCFGHDDVFDPWYPLGTANMLQVLHMGLHVCQ
                     LMGYGQINDGLNLITHHSARTLNLQDYGIAAGNSANLIILPAENGFDALRRQVPVRYSV
                     RGGKVIASTQPAQTTVYLEQPEAIDYKR"
     CDS             5179..5244
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             5578..5883
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     rep_origin      complement(5948..6536)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6028..6047)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6707..7498,7499..7567))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     7330..7349
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7568..7672)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             8013..8318
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
ORIGIN
        1 atggttagta agggtgagga agacaatatg gcaagcctgc cggcaacgca cgaactgcat
       61 atctttggtt cgatcaacgg cgtggacttt gacatggtcg gccaggggac tggtaacccg
      121 aacgatggct acgaggaact gaacctgaaa tcgacaaaag gcgatctgca attctcgcca
      181 tggattctcg tgccgcatat tgggtacggc tttcaccagt atttgccgta tccagatggc
      241 atgagtccgt ttcaagccgc tatggtcgat ggtagcggct atcaggtgca tcgcactatg
      301 cagtttgaag acggcgccag tttgacagtc aattaccgtt acacttatga aggctcgcat
      361 attaaaggtg aggcgcaagt taagggtacc gggtttcccg ccgacggtcc cgtgatgact
      421 aatagcctga ccgcggctga ttggtgccgt agcaaaaaaa cctacccgaa cgacaaaacc
      481 atcatttcca cgtttaaatg gagctatact acaggtaacg ggaagcgcta tcgctcgacg
      541 gcgcgcacta catacacgtt cgcgaaaccg atggccgcga attacctcaa aaaccagccg
      601 atgtatgtgt ttcgtaaaac cgagctgaaa catagcaaga cagaactgaa ctttaaagag
      661 tggcagaaag cattcacaga cgtcatgggc atggatgaac tgtacaaggg cggtggcgat
      721 tacaaagatc acgatggcga ttacaaagat cacgacatcg actataagga tgacgatgat
      781 aagtaggctt caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
      841 tttttgtcgg gaacgctctc tactagagtc acactggctc accttcgggt gggcctttct
      901 gcgggaattc gattgatccg tcgacctgca gggggggggg ggaaagccac gttgtgtctc
      961 aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt
     1021 ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtctt
     1081 gctcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
     1141 gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc
     1201 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     1261 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     1321 ctcctgatga tgcatggtta ctcaccactg cgatccccgg gaaaacagca ttccaggtat
     1381 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     1441 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     1501 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     1561 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taagcttttg ccattctcac
     1621 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     1681 aattaatagg ttgtattgat gttggacgcg tcggaatcgc agaccgatac caggatcttg
     1741 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     1801 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     1861 ttttctaatc agaattggtt aattggttgt aacactggca gagcattacg ctgacttgac
     1921 gggacggcgg ctttgttgaa taaatcgaac ttttgctgag ttgaaggatc agatcacgca
     1981 tcttcccgac aacgcagacc gttccgtggc aaagcaaaag ttcaaaatca ccaactggtc
     2041 cacctacaac aaagctctca tcaaccgtgg ctccctcact ttctggctgg atgatggggc
     2101 gattcaggcc tggtatgagc cagcaacacc ttcttcacga ggcagacctc agcgctcctc
     2161 caccgctgca gttcacttac accgcttctc aacccggtac gcaccagaaa atcattgata
     2221 tggccatgaa tggcgttgga tgccgggcaa cagcccgcat tatgggcgtt ggcctcaaca
     2281 cgattttacg tcacttaaaa aactcaggcc gcagtcggta acctcgcgca tacagccggg
     2341 cagtgacgtc atcgtctgcg cggaaatgga cgaacagtgg ggctatgtcg gggctaaatc
     2401 gcgccagcgc tggctgtttt acgcgtatga cagtctccgg aagacggttg ttgcgcacgt
     2461 attcggtgaa cgcactatgg cgacgctggg gcgtcttatg agcctgctgt caccctttga
     2521 cgtggtgata tggatgacgg atggctggcc gctgtatgaa tcccgcctga agggaaagct
     2581 gcacgtaatc agcaagcgat atacgcagcg aattgagcgg cataacctga atctgaggca
     2641 gcacctggca cggctgggac ggaagtcgct gtcgttctca aaatcggtgg agctgcatga
     2701 caaagtcatc gggcattatc tgaacataaa acactatcaa taagttggaa tcattaccaa
     2761 aaggttagga atacggttag ccatttgcct gcttttatat agttcatatg ggattcacct
     2821 ttatgttgat aagaaataaa agaaaatgcc aataggatat cggcattttc ttttgcgttt
     2881 tcaacgtttg taatcgatgg cttctggctg ctccagatat acggtggttt gtgccggttg
     2941 tgtgctggca atcaccttgc cgccacgtac cgaataacgt accggaacct gacggcgcag
     3001 cgcatcaaac ccattttcag ccggcaggat aatcaggttg gcgctgtttc cggcggcaat
     3061 gccgtaatcc tgcaaattca acgtccttgc gctgtggtgg gtgattaaat tcaggccatc
     3121 gttaatctgc ccgtagccca tcaactggca aacatgcagc cccatatgca gcacttgcag
     3181 catattcgcc gttcccagcg gataccacgg atcgaagaca tcatcgtgac caaagcagac
     3241 gttaatgccc gattccagca tctctttaac gcgcgtgatg ccgcgacgtt ttggatacgt
     3301 atcgaaacgt ccttgcagat gaatattgac cagcgggttg gcgacaaagt taataccgga
     3361 cattttcagc aagcggaaca ggcgtgaggt atacgccccg ttataggagt gcattgccgt
     3421 ggtgtggctg gcggtgacgc gcgcgcccat gccttcatgg tgcgccaggg cagcaacggt
     3481 ttcgacaaag cgcgactgct cgtcatcgat ctcatcacag tgaacgtcga tgagacggtc
     3541 gtatttttgc gccagggcga aggttttatg cagcgattcc acgccgtatt cacgggtaaa
     3601 ttcaaaatgc ggaatcgccc ccactacatc tgcccctaag cgtaacgcct cttccagcaa
     3661 cgcttcaccg ttgggatacg acaaaatccc ttcctgaggg aaggcgacga tttgcagatc
     3721 aatccacggc gcgacttcct gcttcacttc cagcattgct ttcagcgcag ttagcgttgc
     3781 atccgaaaca tcgacatggg tacgcacatg ctgaatgccg ttggcaatct gccatttcag
     3841 cgtttgccat gcgcgttgtt tcacatcgtc atgggttaat aacgctttgc gctcggccca
     3901 gcgttcaatg ccttcaaaca gcgtgccgga ctgattccag ttcggttgtc cggcggtttg
     3961 cgtggtgtcc aggtgaatat gtggctccac aaacggcggt ataactaaac cttgttcggc
     4021 atccaggctg ttttcagtta tgggcatcac gccggattgc gcatcaatgg cgctgatttt
     4081 tccgtcctgc agatgaatct gccacagccc ctcttcgcct ggtaaccggg cgttaataat
     4141 tgtttgtaaa gcgttattcg acatcgttca tgtctccttt tttatgtact gtgttagcgg
     4201 tctgcttctt ccagccctcc tgtttgaaga tggcaagtta gttacgcaca ataaaaaaag
     4261 acctaaaata tgtaaggggt gacgccaaag tatacacttt gccctttaca cattttaggt
     4321 cttgcctgct ttatcagtaa caaacccgcg cgatttactt ttcgacctca ttctattaga
     4381 ttctcgtttg gattgcaact ggtctatttt cctcttttgt ttgatagaaa atcataaaag
     4441 gatttgcaga ctacgggcct aaaggttagt aagggtgagg aagacaatat ggcaagcctg
     4501 ccggcaacgc acgaactgca tatctttggt tcgatcaacg gcgtggactt tgacatggtc
     4561 ggccagggga ctggtaaccc gaacgatggc tacgaggaac tgaacctgaa atcgacaaaa
     4621 ggcgatctgc aattctcgcc atggattctc gtgccgcata ttgggtacgg ctttcaccag
     4681 tatttgccgt atccagatgg catgagtccg tttcaagccg ctatggtcga tggtagcggc
     4741 tatcaggtgc atcgcactat gcagtttgaa gacggcgcca gtttgacagt caattaccgt
     4801 tacacttatg aaggctcgca tattaaaggt gaggcgcaag ttaagggtac cgggtttccc
     4861 gccgacggtc ccgtgatgac taatagcctg accgcggctg attggtgccg tagcaaaaaa
     4921 acctacccga acgacaaaac catcatttcc acgtttaaat ggagctatac tacaggtaac
     4981 gggaagcgct atcgctcgac ggcgcgcact acatacacgt tcgcgaaacc gatggccgcg
     5041 aattacctca aaaaccagcc gatgtatgtg tttcgtaaaa ccgagctgaa acatagcaag
     5101 acagaactga actttaaaga gtggcagaaa gcattcacag acgtcatggg catggatgaa
     5161 ctgtacaagg gcggtggcga ttacaaagat cacgatggcg attacaaaga tcacgacatc
     5221 gactataagg atgacgatga taagtagtga agagctaaaa gccagataac agtatgcgta
     5281 tttgcgcgct gatttttgcg gtataagaat atatactgat atgtataccc gaagtatgtc
     5341 aaaaagaggt atgctatgaa gcagcgtatt acagtgacag ttgacagcga cagctatcag
     5401 ttgctcaagg catatatgat gtcaatatct ccggtctggt aagcacaacc atgcagaatg
     5461 aagcccgtcg tctgcgtgcc gaacgctgga aagcggaaaa tcaggaaggg atggctgagg
     5521 tcgcccggtt tattgaaatg aacggctctt ttgctgacga gaacaggggc tggtgaaatg
     5581 cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt ggatgtacag
     5641 agtgatatta ttgacacgcc cgggcgacgg atggtgatcc ccctggccag tgcacgtctg
     5701 ctgtcagata aagtcccccg tgaactttac ccggtggtgc atatcgggga tgaaagctgg
     5761 cgcatgatga ccaccgatat ggccagtgtg ccggtgtccg ttatcgggga agaagtggct
     5821 gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt ctggggaata
     5881 taaatgtcag gctcccttat acacagccag tctgcaggtc gaccatagtg ctcttcactt
     5941 gagactcttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     6001 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     6061 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6121 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     6181 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6241 ccggtaacta tcgtcttgag cccaacccgg taagacacga cttatcgcca ctggcagcag
     6301 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6361 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     6421 cagttacctt cggaaaaaga gttggtagct cttgatccga caaacaaacc accgctggta
     6481 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6541 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6601 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     6661 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     6721 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctggctcc
     6781 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6841 taccgcgtga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6901 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6961 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     7021 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     7081 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     7141 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     7201 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     7261 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     7321 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     7381 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7441 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     7501 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     7561 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     7621 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cggagtcaga
     7681 cttgaagagc taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata
     7741 agaatatata ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc
     7801 gtattacagt gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa
     7861 tatctccggt ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg
     7921 ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg
     7981 ctcttttgct gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa
     8041 gagagagccg ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc
     8101 gacggatggt gatccccctg gccagtgcac gtctgctgtc agataaagtc ccccgtgaac
     8161 tttacccggt ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca
     8221 gtgtgccggt gtccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca
     8281 tcaaaaacgc cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca
     8341 gccagtctgc aggtcgacca tagtgctctt caggcgatct gggtggcagc ggcggccgc
//
