LOCUS       sequence-8825-ex        5443 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 5443)
  AUTHORS   Harvey CD, Yasuda R, Zhong H, Svoboda K
  TITLE     The spread of Ras activity triggered by activation of a single 
            dendritic spine.
  JOURNAL   Science. 2008 Jul 4. 321(5885):136-40.
  PUBMED    18556515
REFERENCE   2  (bases 1 to 5443)
  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. 
            2008 Jul 4. 321(5885):136-40."
COMMENT     Sequence Label: mEGFP
FEATURES             Location/Qualifiers
     source          1..5443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      934..943
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(940..942,943..945,946..1659)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(985..1006)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1246..1265)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1593..1614
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      1688..1697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1701..1835
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1738..1757)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1792..1811
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        1908..2265
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1908..2099
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1931..1951)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2116..2251
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2285..2302)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2285..2301)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2294..2316)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2514..2969
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2601..2620)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2811..2832
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2986..3005)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3140..3162
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3215..3233)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3301..3405
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3406..3474,3475..4266)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3624..3643)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4437..5025
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4926..4945
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5179..5196
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(5349..5354,5355..5372,5373..5379)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5387..5403
                     /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)."
     primer_bind     5392..5414
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5411..5427
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5411..5427
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgtaccg gtcgccacca tggtgagcaa gggcgaggag
      961 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1021 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1081 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1141 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1201 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1261 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1321 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1381 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1441 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1501 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1561 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1621 gccgggatca ctctcggcat ggacgagctg tacaagtaat cctctagagt cgacctgcag
     1681 aagcttggcc gccatggccc aacttgttta ttgcagctta taatggttac aaataaagca
     1741 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     1801 ccaaactcat caatgtatct tatcatgtct ggatcgatcg ggaattaatt cggcgcagca
     1861 ccatggcctg aaataacctc tgaaagagga acttggttag gtaccttctg aggcggaaag
     1921 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     1981 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     2041 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     2101 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     2161 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     2221 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct gttaacagct
     2281 tggcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     2341 atcgccttgc agcacatccc cccttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     2401 atcgcccttc ccaacagttg cgtagcctga atggcgaatg gcgcctgatg cggtattttc
     2461 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
     2521 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     2581 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     2641 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     2701 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     2761 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     2821 gttccaaact ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat
     2881 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     2941 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
     3001 tgccgcatag ttaagccaac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
     3061 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     3121 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     3181 aaacgcgcga ggcagtattc ttgaagacga aagggcctcg tgatacgcct atttttatag
     3241 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     3301 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     3361 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     3421 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     3481 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     3541 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     3601 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtga tgacgccggg
     3661 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     3721 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     3781 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     3841 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     3901 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgccagc agcaatggca
     3961 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     4021 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     4081 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     4141 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     4201 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     4261 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     4321 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     4381 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     4441 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     4501 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     4561 agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag
     4621 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     4681 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     4741 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     4801 accgaactga gatacctaca gcgtgagcat tgagaaagcg ccacgcttcc cgaagggaga
     4861 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     4921 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     4981 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     5041 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     5101 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     5161 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc
     5221 aaaccgcctc tccccgcgcg ttggccgatt cattaatcca gctggcacga caggtttccc
     5281 gactggaaag cgggcagtga gcgcaacgca attaatgtga gttacctcac tcattaggca
     5341 ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa
     5401 caatttcaca caggaaacag ctatgaccat gattacgaat taa
//
