LOCUS Exported 5282 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS mEGFP-HRas G12V 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 May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5282 /organism="synthetic DNA construct" /mol_type="other DNA" 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 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1079..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 1808..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 3595..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 //