LOCUS       sequence-66948-e        6374 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 6374)
  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 6374)
  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-mEGFP
FEATURES             Location/Qualifiers
     source          1..6374
                     /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             join(2320..2322,2323..2325,2326..3039)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLK
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2365..2386)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2626..2645)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    3162..3283
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3199..3218)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3253..3272
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3290..3745)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3427..3448)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3639..3658
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3772..3876
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3878..4235
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3878..4069
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3901..3921)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4086..4221
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4270..5064
                     /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             4270..5064
                     /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(4324..4343)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4934..4953
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5252..5271)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5296..5343
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5672..6260
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6161..6180
                     /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 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     2401 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     2461 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     2521 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     2581 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     2641 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     2701 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     2761 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     2821 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     2881 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc
     2941 ctgagcaaag accccaacga gaagcgcgat cacatggtcc ttaaggagtt cgtgaccgcc
     3001 gccgggatca ctctcggcat ggacgagctg tacaagtaaa gcggccgcga ctctagatca
     3061 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     3121 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     3181 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3241 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     3301 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     3361 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     3421 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     3481 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     3541 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     3601 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     3661 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     3721 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     3781 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     3841 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     3901 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3961 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     4021 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     4081 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     4141 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     4201 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     4261 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     4321 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4381 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4441 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4501 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4561 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4621 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4681 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4741 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     4801 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4861 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4921 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4981 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     5041 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     5101 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     5161 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     5221 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     5281 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     5341 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     5401 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     5461 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     5521 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5581 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5641 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5701 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5761 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     5821 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5881 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5941 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6001 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6061 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6121 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     6181 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     6241 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     6301 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     6361 ttaccgccat gcat
//
