LOCUS       sequence-66949-e        6365 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dominant-negative EHD2 mutant.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6365)
  AUTHORS   Stoeber M, Stoeck IK, Hanni C, Bleck CK, Balistreri G, Helenius A
  TITLE     Oligomers of the ATPase EHD2 confine caveolae to the plasma membrane
            through association with actin.
  JOURNAL   EMBO J. 2012 May 16;31(10):2350-64. doi: 10.1038/emboj.2012.98. Epub
            2012 Apr 13.
  PUBMED    22505029
REFERENCE   2  (bases 1 to 6365)
  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/emboj.2012.98"; journalName: "EMBO J"; date: "2012-05-16- 
            16"; volume: "31"; issue: "10"; pages: "2350-64"
COMMENT     Sequence Label: EHD2-T72A-mCherry
FEATURES             Location/Qualifiers
     source          1..6365
                     /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"
     protein_bind    628..652
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2282..2306)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      2314..2323
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2320..3030
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2465..2483)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2733..2752
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3153..3274
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3190..3209)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3244..3263
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3281..3736)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3418..3439)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3630..3649
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3763..3867
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3869..4226
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3869..4060
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3892..3912)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4077..4212
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4261..5055
                     /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             4261..5055
                     /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(4315..4334)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4925..4944
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5243..5262)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5287..5334
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5663..6251
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6152..6171
                     /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 ccggactcag atctcgagct caagcttaca agtttgtaca aaaaagttgg catgttcagc
      661 tggctgaagc ggggcggggc acggggccag cagcccgagg ccatccgcac ggtgacctcg
      721 gccctcaagg agctgtaccg cacgaagctg ctgccgctgg aggagcacta ccgctttggg
      781 gccttccact cgccggccct ggaggacgca gacttcgacg gcaagcccat ggtgctggtg
      841 gccggccagt acagcacggg caaggccagc ttcatccagt acctgctgga gcaggaggtg
      901 cccggctccc gcgtggggcc tgagcccacc accgactgct ttgtggccgt catgcacggg
      961 gacactgagg gcaccgtgcc cggcaacgcc ctcgtcgtgg acccggacaa gcccttccgc
     1021 aaactcaacc ctttcggaaa caccttcctc aacaggttca tgtgtgccca gctccctaat
     1081 caggtcctgg agagcatcag catcatcgac accccgggta tcctgtcggg tgccaagcag
     1141 agagtgagcc gcggctacga cttcccggcc gtgctgcgct ggttcgcgga gcgcgtggac
     1201 ctcatcatcc tgctctttga tgcgcacaag ctggagatct cggacgagtt ctcagaggcc
     1261 atcggcgcgt tgcggggcca tgaggacaag atccgcgtgg tgctcaacaa ggccgacatg
     1321 gtggagacgc agcagcttat gcgcgtctac ggcgcgctca tgtgggcgct gggcaaggtg
     1381 gtgggcacgc ccgaggtgct gcgcgtctac atcggctcct tctggtccca gcccctcctc
     1441 gtgcccgaca accggcgcct cttcgagctg gaggagcagg acctcttccg cgacatccag
     1501 ggcctgcccc ggcacgcagc cttgcgcaag ctcaacgacc tggtgaagag ggcccggctg
     1561 gtgcgagttc acgcttacat catcagctac ctgaagaagg agatgccctc tgtgtttggg
     1621 aaggagaaca agaagaagca gctgatcctc aaactgcccg tcatctttgc gaagattcag
     1681 ctggaacatc acatctcccc tggggacttt cctgattgcc agaaaatgca ggagctgctg
     1741 atggcgcatg acttcaccaa gtttcactcg ctgaagccga agctgctgga ggcactggac
     1801 gagatgctga cgcacgacat cgccaagctc atgcccctgc tgcggcagga ggagctggag
     1861 agcaccgagg tgggcgtgca ggggggcgct tttgagggca cccacatggg cccgtttgtg
     1921 gagcggggac ctgacgaggc catggaggac ggcgaggagg gctcggacga cgaggccgag
     1981 tgggtggtga ccaaggacaa gtccaaatac gacgagatct tctacaacct ggcgcctgcc
     2041 gacggcaagc tgagcggctc caaggccaag acctggatgg tggggaccaa gctccccaac
     2101 tcagtgctgg ggcgcatctg gaagctcagc gatgtggacc gcgacggcat gctggatgat
     2161 gaagagttcg cgctggccag ccacctcatc gaggccaagc tggaaggcca cgggctgccc
     2221 gccaacctgc cccgtcgcct ggtgccaccc tccaagcgac gccacaaggg ctccgccgag
     2281 tgcccaactt tcttgtacaa agtggtaccg gtcgccacca tggtgagcaa gggcgaggag
     2341 gataacatgg ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg
     2401 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag
     2461 accgccaagc tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc
     2521 cctcagttca tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac
     2581 ttgaagctgt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc
     2641 ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg
     2701 aagctgcgcg gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc
     2761 tgggaggcct cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag
     2821 cagaggctga agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag
     2881 gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc
     2941 tcccacaacg aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc
     3001 accggcggca tggacgagct gtacaagtaa agcggccgcg actctagatc ataatcagcc
     3061 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
     3121 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     3181 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     3241 gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag cgttaatatt
     3301 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     3361 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     3421 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     3481 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     3541 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     3601 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     3661 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     3721 ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     3781 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     3841 cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt
     3901 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3961 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     4021 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     4081 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     4141 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     4201 tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc
     4261 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     4321 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     4381 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     4441 caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     4501 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     4561 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     4621 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     4681 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     4741 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac
     4801 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     4861 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     4921 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     4981 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     5041 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     5101 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     5161 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     5221 cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg
     5281 ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac
     5341 gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc
     5401 aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag
     5461 ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac
     5521 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     5581 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     5641 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     5701 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     5761 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     5821 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     5881 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     5941 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6001 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6061 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6121 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6181 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     6241 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     6301 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca
     6361 tgcat
//
