LOCUS       sequence-8823-ex        5348 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 5348)
  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 5348)
  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: paGFP-HRas G12V
FEATURES             Location/Qualifiers
     source          1..5348
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     718..742
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1034..1052
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    1052..1132
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      1134..1143
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1140..1142,1143..1145,1146..1856)
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1185..1206)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1446..1465)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1793..1814
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(1869..1901,1902..1904,1905..2438)
                     /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    2462..2583
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2473..2492)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2527..2546
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      2764..3219
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2851..2870)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3061..3082
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3286..3305)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3405..3427
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3465..3483)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3551..3655
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3656..3724,3725..4516)
                     /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(3874..3893)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4687..5275
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5176..5195
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tcgtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt acttaatacg actcactata ggctagcgct accggactca gatctcgagc
     1081 tcaagcttcg aattctgcag tcgacggtac cgcgggcccg ggatccaccg gtcgccacca
     1141 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1201 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1261 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1321 tcgtgaccac cttcagctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     1381 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1441 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1501 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1561 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     1621 gcatcaaggc caacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     1681 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1741 acctgagcca ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1801 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtccg
     1861 gactcgagat gacggaatat aagctggtgg tggtgggcgc cgtcggtgtg ggcaagagtg
     1921 cgctgaccat ccagctgatc cagaaccatt ttgtggacga atacgacccc actatagagg
     1981 attcctaccg gaagcaggtg gtcattgatg gggagacgtg cctgttggac atcctggata
     2041 ccgccggcca ggaggagtac agcgccatgc gggaccagta catgcgcacc ggggagggct
     2101 tcctgtgtgt gtttgccatc aacaacacca agtcttttga ggacatccac cagtacaggg
     2161 agcagatcaa acgggtgaag gactcggatg acgtgcccat ggtgctggtg gggaacaagt
     2221 gtgacctggc tgcacgcact gtggaatctc ggcaggctca ggacctcgcc cgaagctacg
     2281 gcatccccta catcgagacc tcggccaaga cccggcaggg agtggaggat gccttctaca
     2341 cgttggtgcg tgagatccgg cagcacaagc tgcggaagct gaaccctcct gatgagagtg
     2401 gccccggctg catgagctgc aagtgtgtgc tctcctgagc ggccgcttcg agcagacatg
     2461 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     2521 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     2581 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt
     2641 ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg ataaggatcc gggctggcgt
     2701 aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa
     2761 tggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     2821 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     2881 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     2941 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     3001 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3061 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     3121 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     3181 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc ctgatgcggt attttctcct
     3241 tacgcatctg tgcggtattt cacaccgcat atggtgcact ctcagtacaa tctgctctga
     3301 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     3361 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     3421 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     3481 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     3541 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3601 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     3661 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     3721 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     3781 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     3841 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     3901 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     3961 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4021 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4081 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4141 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4201 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4261 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4321 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4381 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4441 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4501 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     4561 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     4621 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     4681 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     4741 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     4801 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     4861 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     4921 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     4981 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5041 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5101 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5161 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5221 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5281 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatggctcg
     5341 acagatct
//
