LOCUS       sequence-9491-ex        6827 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 6827)
  AUTHORS   Inoue T, Heo WD, Grimley JS, Wandless TJ, Meyer T
  TITLE     An inducible translocation strategy to rapidly activate and inhibit 
            small GTPase signaling pathways.
  JOURNAL   Nat Methods. 2005 Jun . 2(6):415-8.
  PUBMED    15908919
REFERENCE   2  (bases 1 to 6827)
  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 
            Methods. 2005 Jun . 2(6):415-8."
COMMENT     Sequence Label: YF-Tiam1
FEATURES             Location/Qualifiers
     source          1..6827
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1342..1662
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     polyA_signal    3615..3736
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3652..3671)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3706..3725
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3743..4198)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3880..3901)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4092..4111
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4225..4329
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4331..4688
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4331..4522
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4354..4374)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4539..4674
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4723..5517
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4723..5517
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4777..4796)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5387..5406
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5705..5724)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5749..5796
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6125..6713
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6614..6633
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccttcggc tacggcctgc agtgcttcgc ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaaga gtgctggtgg tggagtgcag gtggaaacca tctccccagg agacgggcgc
     1381 accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct tgaagatgga
     1441 aagaaatttg attcctcccg ggacagaaac aagcccttta agtttatgct aggcaagcag
     1501 gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca gagagccaaa
     1561 ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat catcccacca
     1621 catgccactc tcgtcttcga tgtggagctt ctaaaactgg aaagtgctgg tggtagtgct
     1681 ggtggtagtg ctggtggtag tgctggtggt agtgctggtg gtcctcgagc aatgaacccc
     1741 tctgaccaga acccatctcc tcaggactcc acggggcctc agctggcgac catgagacaa
     1801 ctctcggatg cagataacgt gcgcaaggtg atctgcgagc tcctggagac ggagcgcacc
     1861 tacgtgaagg atttaaactg tcttatggag agatacctaa agcctcttca aaaagaaact
     1921 tttctcaccc aggatgagct tgacgtgctt tttggaaatt taacggaaat ggtagagttt
     1981 caagtagaat tccttaaaac tctagaagat ggagtgagac tggtacctga tttggaaaag
     2041 cttgagaagg ttgatcaatt taagaaagtg ctgttctctc tggggggatc attcctgtat
     2101 tatgctgacc gcttcaagct ctacagtgcc ttctgcgcca tccacacaaa agttcccaag
     2161 gtcctggtga aagccaagac agacacggct ttcaaggcat tcttggatgc ccagaacccg
     2221 aagcagcagc actcatccac gctggagtcg tacctcatca agcccatcca gaggatcctc
     2281 aagtacccac ttctgctcag ggagctgttc gccctgaccg atgcggagag cgaggagcac
     2341 taccacctgg acgtggccat caagaccatg aacaaggttg ccagtcacat caatgagatg
     2401 cagaaaatcc atgaagagtt tggggctgtg tttgaccagc tgattgctga acagactggt
     2461 gagaaaaaag aggttgcaga tctgagcatg ggagacctgc ttttgcacac taccgtgatc
     2521 tggctgaacc cgccggcctc gctgggcaag tggaaaaagg aaccagagtt ggcagcattc
     2581 gtcttcaaaa ctgctgtggt ccttgtgtat aaagatggtt ccaaacagaa gaagaaactt
     2641 gtaggatctc acaggctttc catttatgag gactgggacc ccttcagatt tcgacacatg
     2701 atccccacgg aagcgctgca ggttcgagct ttggcgagtg cagatgcaga ggcaaatgcc
     2761 gtgtgtgaaa ttgtccatgt aaaatccgag tctgaaggga ggccggagag ggtctttcac
     2821 ttgtgctgca gctccccaga gagccgaaag gatttcctaa aggctgtgca ttcaatcctg
     2881 cgtgataagc acagaagaca gctcctcaaa accgagagcc ttccctcatc ccagcaatat
     2941 gtcccttttg gaggcaaaag attgtgtgca ctgaaggggg ccaggccggc catgagcagg
     3001 gcagtgtctg ccccaagcaa gtctcttggg aggaggaggc ggcggctggc tcgaaacagg
     3061 tttaccattg attctgatgc cgtctccgca agcagcccgg agaaagagtc ccagcagccc
     3121 cccggtggtg gggacactga ccgatgggta gaggagcagt ttgatcttgc tcagtatgag
     3181 gagcaagatg acatcaagga gacagacatc ctcagtgacg atgatgagtt ctgtgagtcc
     3241 gtgaagggtg cctcagtgga cagagacctg caggagcggc ttcaggccac ctccatcagt
     3301 cagcgggaaa gaggccggaa aaccctggat agtcacgcgt cccgcatggc acagctcaag
     3361 aagcaagctg ccctgtcggg gatcaatgga ggcctggaga gcgcaagcga ggaagtcatt
     3421 tgggttaggc gtgaagactt tgccccctcc aggaaactga acactgagat ctgaccgcgg
     3481 gcccgggatc caccggatct agataactga tcataatcag ccataccaca tttgtagagg
     3541 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     3601 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     3661 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     3721 tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     3781 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     3841 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     3901 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     3961 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     4021 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     4081 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     4141 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     4201 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     4261 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     4321 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     4381 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4441 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     4501 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     4561 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     4621 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     4681 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     4741 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     4801 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     4861 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg
     4921 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     4981 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     5041 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     5101 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     5161 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     5221 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg
     5281 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     5341 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     5401 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     5461 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     5521 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     5581 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     5641 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc
     5701 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa
     5761 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg
     5821 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt
     5881 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg
     5941 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac
     6001 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     6061 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     6121 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     6181 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     6241 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     6301 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     6361 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     6421 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     6481 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     6541 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     6601 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     6661 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     6721 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     6781 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
