LOCUS       sequence-12153-e        8127 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 8127)
  AUTHORS   Karginov AV, Ding F, Kota P, Dokholyan NV, Hahn KM
  TITLE     Engineered allosteric activation of kinases in living cells.
  JOURNAL   Nat Biotechnol. 2010 Jul;28(7):743-7. doi: 10.1038/nbt.1639. Epub 
            2010 Jun 27.
  PUBMED    20581846
REFERENCE   2  (bases 1 to 8127)
  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"; doi: "10.1038/nbt.1639"; 
            journalName: "Nat Biotechnol"; date: "2010-07"; volume: "28"; issue:
            "7"; pages: "743-7"
COMMENT     Sequence Label: pEGFP-RapR-FAK-YM-Y397F
FEATURES             Location/Qualifiers
     source          1..8127
                     /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 GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     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"
     polyA_signal    4915..5036
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4952..4971)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5006..5025
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5043..5498)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5180..5201)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5392..5411
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5525..5629
                     /gene="bla"
                     /label=AmpR promoter
     promoter        5631..5988
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5631..5822
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5654..5674)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5839..5974
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6023..6817
                     /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             6023..6817
                     /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(6077..6096)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6687..6706
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7005..7024)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7049..7096
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7425..8013
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7914..7933
                     /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 caccctgacc tacggcgtgc agtgcttcag 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 cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctggaatg gcagctgctt atcttgaccc aaacttgaat
     1381 cacacaccaa gttcgagtac taagactcac ctgggtactg gcatggaacg gtcccctggt
     1441 gcaatggaac gagtattaaa ggtctttcat cattttgaaa gcagtagtga gccaaccacc
     1501 tgggccagta ttatcaggca tggagatgct actgatgtca ggggcatcat tcagaagata
     1561 gtggacagtc acaaagtaaa gcacgtggcc tgctatggat ttcgcctcag tcacctgcga
     1621 tcggaggagg tgcactggct gcacgtggac atgggtgtct ccagtgtgag ggagaaatat
     1681 gaactcgccc acccaccaga ggagtggaaa tatgagttga gaattcgtta cttgccaaaa
     1741 ggatttctaa accagtttac tgaagataag ccaacattga atttcttcta tcaacaggtg
     1801 aagagtgact acatgcaaga aatagctgat caagtagacc aagaaatagc tttgaagttg
     1861 ggttgtttgg aaattaggcg atcctattgg gagatgaggg gtaatgccct agagaagaag
     1921 tccaactatg aagtattaga aaaagatgtt ggtttaaagc gattttttcc taagagttta
     1981 ctggattctg tcaaggccaa aacactaaga aagctgatcc agcaaacatt caggcagttt
     2041 gccaacctta atagagaaga aagcatcctg aaattctttg agattctttc tcctgtgtac
     2101 agatttgaca aagagtgctt caagtgtgcc cttgggtcaa gttggatcat ttctgtggaa
     2161 ttggcaatcg gcccagaaga agggatcagt tacctgacag acaaaggctg caatcccaca
     2221 cacctagcag actttaacca agtgcagacc atccagtact caaacagtga agacaaagac
     2281 aggaaaggaa tgctacaact caaaatcgct ggtgctcctg agcctctgac cgtgacggca
     2341 ccatccctaa ccattgccga gaacatggcc gacctgattg atggatactg ccggctggtg
     2401 aatggagcca cacaatcttt catcatcaga ccccagaaag aaggtgaacg ggcgttgcca
     2461 tcaataccaa agttggccaa cagtgaaaag caaggcatgc ggacacatgc agtctctgtg
     2521 tcagagacag atgactatgc agagatcatc gatgaggaag acacatacac catgccctcg
     2581 accagggatt atgagattca gagagaaaga atagaacttg gacgctgtat tggagaaggt
     2641 cagtttggag atgtacatca aggcgtgtac ggcccaggaa cctgcgtggt gcactacacc
     2701 gggatgcttg aagatggaaa gaaatttgat tcctcccggg acagaaacaa gccctttaag
     2761 tttatgctag gcaagcagga ggtgatccga ggctgggaag aaggggttgc ccagatgagt
     2821 gtgggtcaga gagccaaact gactatatct ccagattatg cctatggtgc cactgggcac
     2881 ccaggcatca tcccaccaca tgccactctc gtcttcgatg tggagcttct aaaactggaa
     2941 ggcccaggtt tggctgttgc aatcaaaaca tgtaaaaact gtacttcgga cagcgtgaga
     3001 gagaagttcc ttcaagaagc cttaacaatg cgtcagtttg accatcctca catcgtgaag
     3061 ctgattggag tcattacaga gaaccctgtc tggataatca tggagttgtg cacacttgga
     3121 gagctgaggt catttttgca agtaagaaaa tacagcttgg acctggcatc tttgatatta
     3181 tatgcctatc agcttagtac agcactcgcg tatctggaga gcaaaagatt tgttcacagg
     3241 gacattgctg ctcggaatgt tctggtgtct tcaaatgatt gtgtaaaatt gggagacttt
     3301 ggattatctc gatatatgga agacagtact tactataaag cttccaaagg aaaattacct
     3361 attaaatgga tggctccaga gtcaatcaat tttcgacgtt ttacctcagc tagtgacgtg
     3421 tggatgtttg gtgtgtgtat gtgggagata ctgatgcatg gcgtgaagcc ttttcaagga
     3481 gtgaagaaca atgatgtgat cggtcgaatt gaaaatgggg aaagattacc aatgcctcca
     3541 aattgtcctc ccaccctcta cagccttatg acgaaatgtt gggcctatga ccccagcagg
     3601 cggcccaggt ttactgaact aaaagctcag ctcagcacaa tcctggagga ggagaaggtg
     3661 cagcaagaag aacggatgag gatggaatcc agaagacagg ctaccgtgtc ctgggactct
     3721 ggagggtctg atgaagcacc acccaagccc agcagacctg gttatcctag cccgagatcc
     3781 agtgaaggat tttatcccag cccacagcac atggtccaga ccaatcacta ccaggtctct
     3841 ggctacccgg gttcccatgg aatcccagcc atggctggca gcatttacca aggtcaggca
     3901 tctcttctgg accagacaga attgtggaat cacagacctc aggagatgtc catgtggcag
     3961 cccagtgtgg aggattcggc agctttggac cttcgaggga tgggacaagt gcttccccct
     4021 cacctgatgg aagagcggct aatccgacag cagcaggaaa tggaagaaga ccagcgctgg
     4081 ctggagaagg aagagcgatt cctgaaacct gatgtgaggc tctctcgagg cagcatcgac
     4141 agggaagacg ggagttttca gggtccgact ggaaaccaac acatctatca gcctgtgggg
     4201 aagccagatc ctgcagctcc accaaagaaa cctcctcgcc ctggagcacc tggccaccta
     4261 agcaacctgt ccagcatcag cagccctgca gacagctaca atgagggtgt caagcttcag
     4321 ccccaggaaa tcagcccccc tcccactgcc aaccttgacc ggtccaatga caaggtatat
     4381 gaaaatgtga caggcctagt gaaggctgtc atcgagatgt ccagcaaaat ccagccagct
     4441 cctccagaag agtacgtccc tatggtgaag gaagttggcc tggccctgcg gaccttacta
     4501 gccacggtgg atgagaccat tcctgctctt ccagccagca ctcatcgaga gatcgagatg
     4561 gcacagaagc tgctgaactc cgacttaggc gagctcatca gcaagatgaa gctggcgcag
     4621 cagtacgtca tgaccagcct gcagcaggag tataagaagc agatgctgac agctgctcac
     4681 gccctggctg tggatgccaa gaacctactt gatgttattg atcaagcaag actgaaaatg
     4741 ctagggcaga cacggccaca ctgagtcgac ggtaccgcgg gcccgggatc caccggatct
     4801 agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     4861 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     4921 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4981 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc
     5041 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     5101 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     5161 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     5221 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     5281 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     5341 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     5401 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     5461 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     5521 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     5581 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     5641 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     5701 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     5761 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     5821 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     5881 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     5941 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     6001 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     6061 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     6121 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     6181 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     6241 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     6301 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     6361 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     6421 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     6481 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     6541 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     6601 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     6661 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     6721 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     6781 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     6841 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     6901 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     6961 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     7021 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     7081 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     7141 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     7201 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     7261 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     7321 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     7381 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     7441 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     7501 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     7561 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     7621 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     7681 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     7741 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     7801 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     7861 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     7921 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     7981 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     8041 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     8101 gtggataacc gtattaccgc catgcat
//
