LOCUS       sequence-159474-        5006 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 5006)
  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 5006)
  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-pGBD
FEATURES             Location/Qualifiers
     source          1..5006
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..978
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1013..1031
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1013..1030
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             join(1068..1070,1071..1073,1074..1784)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1068..1070,1071..1073,1074..1784)
                     /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(1113..1134)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1374..1393)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1721..1742
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2036..2170
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2060..2079)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2114..2133
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2283..2303)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2283..2301)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2322..2338)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2322..2338)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2335..2357)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2346..2362
                     /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(2370..2376,2377..2394,2395..2400))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2415..2436
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2553..2570)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2724..3312)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2804..2823)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3483..4274,4275..4343))
                     /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     4106..4125
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4344..4448)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4474..4929)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4611..4632)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4823..4842
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
       61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
      121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
      181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
      241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
      301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
      361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
      421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
      481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
      541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
      601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
      661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
      721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
      781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
      841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
      901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
      961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct tgatttaggt
     1021 gacactatag aatacaagct acttgttctt tttgcaggat ccaaaccatg gtgagcaagg
     1081 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     1141 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     1201 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     1261 tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
     1321 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     1381 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     1441 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     1501 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
     1561 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     1621 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
     1681 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1741 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtccgga aaagagcgcc
     1801 cagagatctc tcttccttca gactttgagc atacgattca tgtgggtttt gatgcagtca
     1861 ccggggaatt cactggaatt cctgagcaat gggcacgact actccaaacc tccaacataa
     1921 caaagctgga acagaagaag aacccacaag ctgttcttga tgttctcaag ttctacgact
     1981 ccaaagaaac ggtcaacaac cagaaatcta gaactatagt gagtcgtatt acgtagatcc
     2041 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     2101 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     2161 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     2221 ggaggttttt taattcgcgg ccgcggcgcc aatgcattgg gcccggtacc cagcttttgt
     2281 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     2341 tgaaattgtt atccgctcac aattccacac aacatacgag ccgggagcat aaagtgtaaa
     2401 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     2461 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     2521 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     2581 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     2641 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2701 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2761 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2821 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2881 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2941 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3001 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3061 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3121 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     3181 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3241 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3301 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3361 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3421 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3481 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     3541 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     3601 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     3661 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     3721 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     3781 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     3841 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     3901 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     3961 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4021 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4081 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4141 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4201 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4261 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4321 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4381 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4441 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     4501 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     4561 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     4621 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4681 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     4741 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     4801 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     4861 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4921 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     4981 cctcttcgct attacgccag tcgacc
//
