LOCUS       sequence-235856-        4665 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 4665)
  AUTHORS   Nalbant P, Hodgson L, Kraynov V, Toutchkine A, Hahn KM
  TITLE     Activation of endogenous Cdc42 visualized in living cells.
  JOURNAL   Science. 2004 Sep 10. 305(5690):1615-9.
  PUBMED    15361624
REFERENCE   2  (bases 1 to 4665)
  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: "Science. 
            2004 Sep 10. 305(5690):1615-9."
COMMENT     Sequence Label: pET23-CBD-(-PP)-EGFP
FEATURES             Location/Qualifiers
     source          1..4665
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     51..70
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        51..69
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             101..123
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             join(493..495,496..498,499..1209)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(493..495,496..498,499..1209)
                     /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(538..559)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(799..818)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1146..1167
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1216..1233
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1286..1304)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1300..1347
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1384..1839
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1471..1490)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1681..1702
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1865..1969
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1970..2038,2039..2830)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2188..2207)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3001..3589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3490..3509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3743..3760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3775..3914
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3840..3859)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4000..4022
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4016..4207)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gcacccgtgg ccaggaccca acgctgcccg agatctcgat cccgcgaaat taatacgact
       61 cactataggg agaccacaac ggtttccctc tagaaataat tttgtttaac tttaagaagg
      121 agatatacca tggacatcca gaaccctgac atcacgagtt cacgataccg tgggctccca
      181 gcacctggac ctagcccagc tgataagaaa cgctcaggga agaagaagat cagcaaagct
      241 gatattggtg cacccagtgg attcaagcat gtcagccacg tggggtggga cccccagaat
      301 ggatttgacg tgaacaacct cgacccagat ctgcggagtc tgtgctccag ggcaggaatc
      361 agcgaggccc agctcaccga cgccgagacc tctaaactta tctacgactt cattgaggac
      421 cagggtgggc tggaggctgt gcggcaggag atgaggcgcc aggagccact tgaattcggt
      481 ggcggtggca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      541 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      601 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      661 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      721 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      781 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      841 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
      901 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
      961 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1021 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1081 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1141 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1201 ctgtacaagc tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga
     1261 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     1321 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat
     1381 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     1441 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     1501 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     1561 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     1621 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     1681 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     1741 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     1801 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     1861 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1921 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1981 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     2041 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     2101 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     2161 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     2221 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     2281 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     2341 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     2401 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     2461 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat
     2521 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     2581 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     2641 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2701 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2761 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2821 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     2881 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2941 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     3001 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     3061 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     3121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
     3181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     3241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     3301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     3661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     3721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat
     3781 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt
     3841 acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact
     3901 gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc
     3961 tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga
     4021 ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt
     4081 cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca
     4141 gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt
     4201 tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga
     4261 aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac
     4321 gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg
     4381 gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc
     4441 ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt
     4501 acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc
     4561 agcagtcgct tcacgttcgc tcgcatatcg gtgattcatt ctgctaacca gtaaggcaac
     4621 cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgc
//
