LOCUS       sequence-326715-        7205 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7205)
  AUTHORS   Miller AL, Bement WM
  TITLE     Regulation of cytokinesis by Rho GTPase flux.
  JOURNAL   Nat Cell Biol. 2009 Jan . 11(1):71-7.
  PUBMED    19060892
REFERENCE   2  (bases 1 to 7205)
  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 Cell 
            Biol. 2009 Jan . 11(1):71-7."
COMMENT     Sequence Label: EMTB-3XGFP
FEATURES             Location/Qualifiers
     source          1..7205
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     35..52
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      100..109
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             109..138
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      958..967
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(964..966,967..969,970..1680)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(964..966,967..969,970..1680)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      1723..1732
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1729..1731,1732..1734,1735..2445)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1729..1731,1732..1734,1735..2445)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      2494..2503
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2500..2502,2503..2505,2506..3219)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2500..2502,2503..2505,2506..3216)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    3257..3391
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3281..3300)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3335..3354
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3504..3524)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3504..3522)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3543..3559)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3543..3559)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3556..3578)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3567..3583
                     /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)."
     promoter        complement(join(3591..3597,3598..3615,3616..3621))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3636..3657
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3774..3791)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3945..4533)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4025..4044)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4704..5495,5496..5564))
                     /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     5327..5346
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5565..5669)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5695..6150)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5832..5853)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6044..6063
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        6221..7205
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
       61 ctacttgttc tttttgcagg atcccatcga ttcgaattcg ccaccatgga gcagaagctc
      121 atctcagaag aagacctcgg cgcgccagtg cgaagcgaaa cagcacccga cagctacaaa
      181 gtgcaagata agaaaaatgc ctccagccgc cctgcctctg caatttcagg acaaaataac
      241 aaccactcag gaaataaacc agaccctccg cctgtgttac gtgttgatga ccggcagcgg
      301 ctggcccggg agcgacgtga ggaacgggag aaacagctag ctgcaagaga aatagtgtgg
      361 ttagaaagag aagagcgagc caggcagcac tacgagaagc acctggaaga gcggaagaag
      421 aggttggagg agcagaggca gaaggaggag cggaggaggg ctgctgtgga ggagaagcgg
      481 aggcagagac ttgaggagga caaagaacgc cacgaagctg ttgtacggcg cacaatggaa
      541 aggagccaga agccaaaaca gaagcataac cgttggtcgt ggggaggctc tctccatggg
      601 agccctagca tccacagtgc agatccagac aggcggtcag tttccaccat gaatctttcg
      661 aaatatgttg atcccgtcat tagcaagcgg ctctcctctt catctgcaac tttactaaat
      721 tctccagata gagctcgccg cctgcagctc agcccatggg agagcagcgt tgttaacaga
      781 ctcctgacgc ccacacattc gttcctggcc agaagtaaaa gcacagctgc cttgtctgga
      841 gaagcagcat cttgcagccc catcatcatg ccctacaaag ctgcacactc tagaaattcg
      901 atggatcgac caaaactctt tgtaacacca cctgagggct cttcggatcc accggtcgcc
      961 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     1021 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
     1081 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     1141 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
     1201 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1261 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1321 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1381 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     1441 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     1501 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1561 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     1621 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1681 gaaagcggcc acggtacccc gcgggcccgg gatccaccgg tcgccaccat ggtgagcaag
     1741 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     1801 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     1861 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     1921 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     1981 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     2041 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     2101 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     2161 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     2221 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     2281 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     2341 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     2401 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagggaag tggaccgcgg
     2461 gatatccctg agggctcttc ggatccaccg gtcgccacca tggtgagcaa gggcgaggag
     2521 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     2581 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     2641 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     2701 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     2761 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     2821 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     2881 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     2941 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     3001 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     3061 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc
     3121 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     3181 gccgggatca ctctcggcat ggacgagctg tacaagtaac tcgagcctct agaactatag
     3241 tgagtcgtat tacgtagatc cagacatgat aagatacatt gatgagtttg gacaaaccac
     3301 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     3361 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
     3421 tcaggttcag ggggaggtgt gggaggtttt ttaattcgcg gccgcggcgc caatgcattg
     3481 ggcccggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     3541 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3601 gccgggagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3661 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3721 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3781 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     3841 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     3901 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     3961 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4021 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4081 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4141 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4201 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4261 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4321 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4381 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4441 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4501 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4561 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4621 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4681 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4741 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4801 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4861 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     4921 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     4981 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5041 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5101 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5161 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5221 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5281 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5341 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5401 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5461 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5521 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5581 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5641 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     5701 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     5761 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     5821 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     5881 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     5941 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     6001 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     6061 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     6121 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
     6181 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gtcgaccgcc aattcaatat
     6241 ggcgtatatg gactcatgcc aattcaatat ggtggatctg gacctgtgcc aattcaatat
     6301 ggcgtatatg gactcgtgcc aattcaatat ggtggatctg gaccccagcc aattcaatat
     6361 ggcggacttg gcaccatgcc aattcaatat ggcggacctg gcactgtgcc aactggggag
     6421 gggtctactt ggcacggtgc caagtttgag gaggggtctt ggccctgtgc caagtccgcc
     6481 atattgaatt ggcatggtgc caataatggc ggccatattg gctatatgcc aggatcaata
     6541 tataggcaat atccaatatg gccctatgcc aatatggcta ttggccaggt tcaatactat
     6601 gtattggccc tatgccatat agtattccat atatgggttt tcctattgac gtagatagcc
     6661 cctcccaatg ggcggtccca tataccatat atggggcttc ctaataccgc ccatagccac
     6721 tcccccattg acgtcaatgg tctctatata tggtctttcc tattgacgtc atatgggcgg
     6781 tcctattgac gtatatggcg cctcccccat tgacgtcaat tacggtaaat ggcccgcctg
     6841 gctcaatgcc cattgacgtc aataggacca cccaccattg acgtcaatgg gatggctcat
     6901 tgcccattca tatccgttct cacgccccct attgacgtca atgacggtaa atggcccact
     6961 tggcagtaca tcaatatcta ttaatagtaa cttggcaagt acattactat tggaagtacg
     7021 ccagggtaca ttggcagtac tcccattgac gtcaatggcg gtaaatggcc cgcgatggct
     7081 gccaagtaca tccccattga cgtcaatggg gaggggcaat gacgcaaatg ggcgttccat
     7141 tgacgtaaat gggcggtagg cgtgcctaat gggaggtcta tataagcaat gctcgtttag
     7201 ggaac
//
