LOCUS       sequence-398136-        6473 bp    DNA     circular SYN 14-APR-2025
DEFINITION  fluorescent protein tagged ezrin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6473)
  AUTHORS   Pacheco J, Cassidy AC, Zewe JP, Wills RC, Hammond GRV
  TITLE     PI(4,5)P2 diffuses freely in the plasma membrane even within 
            high-density effector protein complexes.
  JOURNAL   J Cell Biol. 2023 Feb 6;222(2):e202204099. doi: 
            10.1083/jcb.202204099. Epub 2022 Nov 23.
  PUBMED    36416724
REFERENCE   2  (bases 1 to 6473)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2023-02-6"; volume: "222(2):e202204099. doi"; pages: 
            " 10.1083/jcb.202204099. Epub 2022 Nov 23"
COMMENT     Sequence Label: EGFP-N-Ezrin
FEATURES             Location/Qualifiers
     source          1..6473
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        52..243
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     200..220
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(245..349)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      376..831
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(463..482)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     673..694
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    838..959
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(849..868)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     903..922
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(1082..1795,1796..1798,1799..1801))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1085..1795,1796..1798,1799..1801))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1127..1148)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1476..1495
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1735..1756
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      1798..1807
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(3652..3855)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3681..3701)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3856..4159
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(4334..4922)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4414..4433)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    5251..5298
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     5323..5342
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(5530..6324)
                     /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             complement(5530..6324)
                     /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(5641..5660)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6251..6270
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(6359..6473,1..243))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(6373..6473,1..35)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 gaatgggcgg aactgggcgg agttaggggc gggatgggcg gagttagggg cgggactatg
       61 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      121 tttccacacc tggttgctga ctaattgaga tgcatgcttt gcatacttct gcctgctggg
      181 gagcctgggg actttccaca ccctaactga cacacattcc acagctggtt ctttccgcct
      241 caggactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
      301 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
      361 gaaaagtgcc acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
      421 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
      481 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
      541 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
      601 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
      661 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
      721 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
      781 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt tacgccttaa
      841 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
      901 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
      961 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt
     1021 aaagcaagta aaacctctac aaatgtggta tggctgatta tgatctagag tcgcggccgc
     1081 tttacttgta cagctcgtcc atgccgagag tgatcccggc ggcggtcacg aactccagca
     1141 ggaccatgtg atcgcgcttc tcgttggggt ctttgctcag ggcggactgg gtgctcaggt
     1201 agtggttgtc gggcagcagc acggggccgt cgccgatggg ggtgttctgc tggtagtggt
     1261 cggcgagctg cacgctgccg tcctcgatgt tgtggcggat cttgaagttc accttgatgc
     1321 cgttcttctg cttgtcggcc atgatataga cgttgtggct gttgtagttg tactccagct
     1381 tgtgccccag gatgttgccg tcctccttga agtcgatgcc cttcagctcg atgcggttca
     1441 ccagggtgtc gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga
     1501 agaagatggt gcgctcctgg acgtagcctt cgggcatggc ggacttgaag aagtcgtgct
     1561 gcttcatgtg gtcggggtag cggctgaagc actgcacgcc gtaggtcagg gtggtcacga
     1621 gggtgggcca gggcacgggc agcttgccgg tggtgcagat gaacttcagg gtcagcttgc
     1681 cgtaggtggc atcgccctcg ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc
     1741 cgtccagctc gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca
     1801 tggtggcgac cggtggatcc cgggcccgcg gtaccgtcga ctgcagaatt cgaagcttca
     1861 tggcctcgaa ctcgtcaatg cgttgcttgg tgttgccctg cctgatttgc cgcagcgtct
     1921 tatacttgtc cctgccttgc cgcatgttct cgttgtggat gatgtcattg tgggtcctct
     1981 tgttctcatc ccgggcctgg gacaactcat tgctcagggt cagcagctgc cgctgcacgc
     2041 gctcattctt ctctgcctct gtgatccgct tctcctcgtt gcggtcatcc aggatgccct
     2101 cactggagag ctcggcactg tagcccatag gctctgctcc ctcgtcctgc agtccctcct
     2161 gcacgtggta attcacaggc tcatacactg ggggcggtgg gggcggtggg gccgtcatca
     2221 ccaggtgcag ctcctctttg gtcttcacca ggtcgtcctg ggcttcttta gcccggtgct
     2281 gccactcttc tacctcgtcc tccttgcgcc tccgcgcctc ctccagcagt gcgatcttgg
     2341 ccgtgtactc tgccagctct gcagccagct gctcctggct ctttatctga tcctgcgcct
     2401 gtctctcgag ttcttccttg gcccgcaggg cggccatgcg gtcggcctcc agacgttcag
     2461 cctcctcctg ggctcgcctc ctctcttcct ccagttggag ggccttctca atctgctcgg
     2521 agagctcttt ctccgccctc ttggtcttct gctcgtagtc ctgcagccga agcatcagct
     2581 cctccttctc ccggagcatc tgctcctttt ctctctccac cgtctctcgc ctcttcttct
     2641 cggtttccaa ctgctgtcgc tctagctgct tctggtgctt ctcctcccga gcctgggcct
     2701 tcatctgctg cacctcgatg gtgtcgggct tcctgcggcg catgtacagc tcatggttcc
     2761 ccatgcagag ctgcaggatc cgcttgttaa ttctcaggcg cggggcgtag aacacaaagt
     2821 caggtgcctt cttgtcgatg ggcttaatga caaacttctt gtcgttgaaa gagatgttcc
     2881 tgatctcact ccaagggaag ccgatctttg gggtcaactt gtcatctttc tcataaatgt
     2941 taagtccaag ggcatcgact ccaagccaaa ggtctgttcc tttcttgttt ttgatctcga
     3001 aatagttgat cccatacatt tccaggtcct gggcaatctt caggtattct agcatagcac
     3061 tgtccttgag catccctcgg tgttccgcgt gccacacctg gatccggtcc tcccactggt
     3121 ccctgctgag cttgtgttgg tccatgactc tctgggggat cagccgctcc gagctgaggt
     3181 acccagactt gtgcatttcc ttgttataat ctccgaactt ggcctgaacg gcataggagc
     3241 ccaggagcac ggctgtctct ggggggcagt agatctcgtc gctgaggatc ccgtccttga
     3301 cttgcaggaa gaagagcttc tgtgtgatgt cctggatgag ttcctcggcc acgtcttcgg
     3361 ggtagaactt ggcccggaat ttaaactgga cagggttctc ctttcggacc tcctgtgcgg
     3421 agaccttttt atcaagtttc agccaggtag gaaatccttt attgtctaca tactggaggc
     3481 cgaagtacca cacttcccgg aggccaatcg tctttaccac ctgatcaaag agctgcttcc
     3541 cggtggtgtt tggctggatg gcaaactcca gctcggcatc catggtggtc acccggacgt
     3601 tgattggctt gggcatggtg gcggctagcg gatctgacgg ttcactaaac cagctctgct
     3661 tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg cggagttgtt
     3721 acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat tgacgtcaat
     3781 ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat tgatgtactg
     3841 ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact gccaagtagg
     3901 aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac cgtcattgac
     3961 gtcaataggg ggcgtacttg gcatatgata cacttgatgt actgccaagt gggcagttta
     4021 ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta ctatgggaac
     4081 atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag gcgggccatt
     4141 taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg accccgtaat
     4201 tgattactat taataactaa tgcatggcgg taatacggtt atccacagaa tcaggggata
     4261 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4321 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4381 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4441 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4501 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4561 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4621 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4681 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4741 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4801 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4861 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4921 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4981 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     5041 aatgaagttt taaatcaatc taaagtatat atgagtaacc tgaggctatg gcagggcctg
     5101 ccgccccgac gttggctgcg agccctgggc cttcacccga acttgggggg tggggtgggg
     5161 aaaaggaaga aacgcgggcg tattggcccc aatggggtct cggtggggta tcgacagagt
     5221 gccagccctg ggaccgaacc ccgcgtttat gaacaaacga cccaacaccg tgcgttttat
     5281 tctgtctttt tattgccgtc atagcgcggg ttccttccgg tattgtctcc ttccgtgttt
     5341 cagttagcct ccccctaggg tgggcgaaga actccagcat gagatccccg cgctggagga
     5401 tcatccagcc ggcgtcccgg aaaacgattc cgaagcccaa cctttcatag aaggcggcgg
     5461 tggaatcgaa atctcgtgat ggcaggttgg gcgtcgcttg gtcggtcatt tcgaacccca
     5521 gagtcccgct cagaagaact cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg
     5581 agcggcgata ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc
     5641 aatatcacgg gtagccaacg ctatgtcctg atagcggtcc gccacaccca gccggccaca
     5701 gtcgatgaat ccagaaaagc ggccattttc caccatgata ttcggcaagc aggcatcgcc
     5761 atgggtcacg acgagatcct cgccgtcggg catgctcgcc ttgagcctgg cgaacagttc
     5821 ggctggcgcg agcccctgat gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc
     5881 catccgagta cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc
     5941 cggatcaagc gtatgcagcc gccgcattgc atcagccatg atggatactt tctcggcagg
     6001 agcaaggtga gatgacagga gatcctgccc cggcacttcg cccaatagca gccagtccct
     6061 tcccgcttca gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg tggccagcca
     6121 cgatagccgc gctgcctcgt cttgcagttc attcagggca ccggacaggt cggtcttgac
     6181 aaaaagaacc gggcgcccct gcgctgacag ccggaacacg gcggcatcag agcagccgat
     6241 tgtctgttgt gcccagtcat agccgaatag cctctccacc caagcggccg gagaacctgc
     6301 gtgcaatcca tcttgttcaa tcatgcgaaa cgatcctcat cctgtctctt gatcgatctt
     6361 tgcaaaagcc taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg
     6421 aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc gga
//
