LOCUS       sequence-324546-        5067 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 5067)
  AUTHORS   Benink HA, Bement WM
  TITLE     Concentric zones of active RhoA and Cdc42 around single cell wounds.
  JOURNAL   J Cell Biol. 2005 Jan 31. 168(3):429-39.
  PUBMED    15684032
REFERENCE   2  (bases 1 to 5067)
  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: "J Cell 
            Biol. 2005 Jan 31. 168(3):429-39."
COMMENT     Sequence Label: GFP-wGBD
FEATURES             Location/Qualifiers
     source          1..5067
                     /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"
     CDS             join(90..92,93..95,96..806)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(90..92,93..95,96..806)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(135..156)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(396..415)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     743..764
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1119..1253
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1143..1162)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1197..1216
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1366..1386)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1366..1384)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1405..1421)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1405..1421)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1418..1440)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1429..1445
                     /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(1453..1459,1460..1477,1478..1483))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1498..1519
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1636..1653)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1807..2395)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1887..1906)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2566..3357,3358..3426))
                     /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     3189..3208
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3427..3531)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3557..4012)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3694..3715)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3906..3925
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4083..5067
                     /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 atccaaacca tggtgagcaa gggcgaggag ctgttcaccg
      121 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt
      181 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca
      241 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt
      301 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
      361 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
      421 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
      481 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
      541 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
      601 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
      661 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag
      721 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
      781 ctctcggcat ggacgagctg tacaagtccg gacctagccc agctgataag aaacgctcag
      841 ggaagaagaa gatcagcaaa gctgatattg gtgcacccag tggattcaag catgtcagcc
      901 acgtggggtg ggacccccag aatggatttg acgtgaacaa cctcgaccca gatctgcgga
      961 gtctgttctc cagggcagga atcagcgagg cccagctcac cgacgccgag acctctaaac
     1021 ttatctacga cttcattgag gaccagggtg ggctggaggc tgtgcggcag gagatgaggc
     1081 gccaggagtt ctagaactat agtgagtcgt attacgtaga tccagacatg ataagataca
     1141 ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa
     1201 tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca
     1261 acaattgcat tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaattcg
     1321 cggccgcggc gccaatgcat tgggcccggt acccagcttt tgttcccttt agtgagggtt
     1381 aattgcgcgc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     1441 cacaattcca cacaacatac gagccgggag cataaagtgt aaagcctggg gtgcctaatg
     1501 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     1561 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     1621 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     1681 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     1741 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1801 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1861 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1921 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1981 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     2041 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2101 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2161 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     2221 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     2281 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2341 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2401 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2461 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2521 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     2581 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     2641 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     2701 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     2761 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     2821 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     2881 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     2941 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     3001 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     3061 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     3121 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     3181 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     3241 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     3301 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     3361 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     3421 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     3481 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     3541 ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt
     3601 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
     3661 aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
     3721 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
     3781 cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     3841 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     3901 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     3961 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg
     4021 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     4081 cagtcgaccg ccaattcaat atggcgtata tggactcatg ccaattcaat atggtggatc
     4141 tggacctgtg ccaattcaat atggcgtata tggactcgtg ccaattcaat atggtggatc
     4201 tggaccccag ccaattcaat atggcggact tggcaccatg ccaattcaat atggcggacc
     4261 tggcactgtg ccaactgggg aggggtctac ttggcacggt gccaagtttg aggaggggtc
     4321 ttggccctgt gccaagtccg ccatattgaa ttggcatggt gccaataatg gcggccatat
     4381 tggctatatg ccaggatcaa tatataggca atatccaata tggccctatg ccaatatggc
     4441 tattggccag gttcaatact atgtattggc cctatgccat atagtattcc atatatgggt
     4501 tttcctattg acgtagatag cccctcccaa tgggcggtcc catataccat atatggggct
     4561 tcctaatacc gcccatagcc actcccccat tgacgtcaat ggtctctata tatggtcttt
     4621 cctattgacg tcatatgggc ggtcctattg acgtatatgg cgcctccccc attgacgtca
     4681 attacggtaa atggcccgcc tggctcaatg cccattgacg tcaataggac cacccaccat
     4741 tgacgtcaat gggatggctc attgcccatt catatccgtt ctcacgcccc ctattgacgt
     4801 caatgacggt aaatggccca cttggcagta catcaatatc tattaatagt aacttggcaa
     4861 gtacattact attggaagta cgccagggta cattggcagt actcccattg acgtcaatgg
     4921 cggtaaatgg cccgcgatgg ctgccaagta catccccatt gacgtcaatg gggaggggca
     4981 atgacgcaaa tgggcgttcc attgacgtaa atgggcggta ggcgtgccta atgggaggtc
     5041 tatataagca atgctcgttt agggaac
//
