LOCUS       sequence-182619-        5282 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 5282)
  AUTHORS   Yasuda R, Harvey CD, Zhong H, Sobczyk A, van Aelst L, Svoboda K
  TITLE     Supersensitive Ras activation in dendrites and spines revealed by 
            two-photon fluorescence lifetime imaging.
  JOURNAL   Nat Neurosci. 2006 Feb . 9(2):283-91.
  PUBMED    16429133
REFERENCE   2  (bases 1 to 5282)
  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 
            Neurosci. 2006 Feb . 9(2):283-91."
COMMENT     Sequence Label: mEGFP-HRas G12V
FEATURES             Location/Qualifiers
     source          1..5282
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        143..522
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        523..726
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     677..697
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     723..747
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          862..994
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     977..996
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1039..1057
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1073..1082
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1079..1081,1082..1084,1085..1795)
                     /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(1124..1145)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1385..1404)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1732..1753
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(1808..1840,1841..1843,1844..2377)
                     /codon_start=1
                     /product="human oncoprotein generated by the G12V mutation 
                     in the small GTPase H-Ras"
                     /label=H-Ras (G12V)
                     /translation="MTEYKLVVVGAVGVGKSALTIQLIQNHFVDEYDPTIEDSYRKQVV
                     IDGETCLLDILDTAGQEEYSAMRDQYMRTGEGFLCVFAINNTKSFEDIHQYREQIKRVK
                     DSDDVPMVLVGNKCDLAARTVESRQAQDLARSYGIPYIETSAKTRQGVEDAFYTLVREI
                     RQHKLRKLNPPDESGPGCMSCKCVLS"
     polyA_signal    2401..2522
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2412..2431)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2466..2485
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      2703..3158
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2790..2809)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3000..3021
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3225..3244)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3344..3366
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3404..3422)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3490..3594
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3595..3663,3664..4455)
                     /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     complement(3813..3832)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4626..5214
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5115..5134
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gatcttcaat attggccatt agccatatta ttcattggtt atatagcata aatcaatatt
       61 ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta tattggctca
      121 tgtccaatat gaccgccatg ttggcattga ttattgacta gttattaata gtaatcaatt
      181 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
      241 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
      301 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
      361 actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc tattgacgtc
      421 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg ggactttcct
      481 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
      541 tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
      601 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaat
      661 aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
      721 agagctcgtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt ttatcacagt
      781 taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag ctgcagaagt
      841 tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta aggagaccaa
      901 tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc acctattggt
      961 cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccagtt caattacagc
     1021 tcttaaggct agagtactta atacgactca ctataggcta gcgctaccgg tcgccaccat
     1081 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1141 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1201 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1261 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1321 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     1381 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     1441 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     1501 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     1561 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     1621 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     1681 cctgagcacc cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     1741 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtccgg
     1801 actcgagatg acggaatata agctggtggt ggtgggcgcc gtcggtgtgg gcaagagtgc
     1861 gctgaccatc cagctgatcc agaaccattt tgtggacgaa tacgacccca ctatagagga
     1921 ttcctaccgg aagcaggtgg tcattgatgg ggagacgtgc ctgttggaca tcctggatac
     1981 cgccggccag gaggagtaca gcgccatgcg ggaccagtac atgcgcaccg gggagggctt
     2041 cctgtgtgtg tttgccatca acaacaccaa gtcttttgag gacatccacc agtacaggga
     2101 gcagatcaaa cgggtgaagg actcggatga cgtgcccatg gtgctggtgg ggaacaagtg
     2161 tgacctggct gcacgcactg tggaatctcg gcaggctcag gacctcgccc gaagctacgg
     2221 catcccctac atcgagacct cggccaagac ccggcaggga gtggaggatg ccttctacac
     2281 gttggtgcgt gagatccggc agcacaagct gcggaagctg aaccctcctg atgagagtgg
     2341 ccccggctgc atgagctgca agtgtgtgct ctcctgagcg gccgcttcga gcagacatga
     2401 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     2461 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     2521 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt
     2581 tttaaagcaa gtaaaacctc tacaaatgtg gtaaaatcga taaggatccg ggctggcgta
     2641 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
     2701 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2761 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2821 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2881 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     2941 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3001 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     3061 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     3121 taacgcgaat tttaacaaaa tattaacgct tacaatttcc tgatgcggta ttttctcctt
     3181 acgcatctgt gcggtatttc acaccgcata tggtgcactc tcagtacaat ctgctctgat
     3241 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     3301 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     3361 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     3421 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     3481 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3541 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     3601 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3661 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3721 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3781 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     3841 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3901 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3961 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4021 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4081 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4141 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4201 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4261 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4321 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     4381 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4441 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4501 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4561 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4621 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4681 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4741 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4801 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4861 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4921 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4981 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5041 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5101 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5161 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5221 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catggctcga
     5281 ca
//
