LOCUS       sequence-384484-        5813 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression vector for GFP tagged NHERF1. Triple PKC site 
            phosphorylation-deficient mutant (S162/339/340A).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5813)
  AUTHORS   Garbett D, LaLonde DP, Bretscher A
  TITLE     The scaffolding protein EBP50 regulates microvillar assembly in a 
            phosphorylation-dependent manner.
  JOURNAL   J Cell Biol. 2010 Oct 18;191(2):397-413. doi: 10.1083/jcb.201004115.
            Epub 2010 Oct 11.
  PUBMED    20937695
REFERENCE   2  (bases 1 to 5813)
  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: 
            "10.1083/jcb.201004115"; journalName: "J Cell Biol"; date: 
            "2010-10-18- 18"; volume: "191"; issue: "2"; pages: "397-413"
COMMENT     Sequence Label: GFP-NHERF1 3xSA
FEATURES             Location/Qualifiers
     source          1..5813
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     164..185
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1499..1620
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1536..1555)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1590..1609
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1627..2082)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1764..1785)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1976..1995
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2109..2213
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2215..2572
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2215..2406
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2238..2258)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2423..2558
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2607..3401
                     /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             2607..3401
                     /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(2661..2680)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3271..3290
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3589..3608)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3633..3680
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4009..4597
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4498..4517
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4772..5075
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5076..5279
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5230..5250
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      5318..5327
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(5324..5326,5327..5329,5330..5813,1..227)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(5324..5326,5327..5329,5330..5813,1..227)
                     /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(5369..5390)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5630..5649)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
       61 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
      121 cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
      181 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtcc ggccggactc
      241 agatcctgat gagcgcggac gcagcggccg gggcgcccct gccccggctc tgctgcctgg
      301 agaagggtcc gaacggctac ggcttccacc tgcacgggga gaagggcaag ttgggccagt
      361 acatccggct ggtggagccc ggctcgccgg ccgagaaggc ggggctgctg gcgggggacc
      421 ggctggtgga ggtgaacggg gagaacgtgg aaaaggagac ccaccagcag gtggtgagcc
      481 gcatccgcgc cgcactcaac gccgtgcgcc tgctggtggt cgaccccgag acggacgagc
      541 agctgcagaa gctcggcgtc caggtccgag aggagctgct gcgcgcccag gaagcgccgg
      601 ggcaggccga gccgccggcc gccgccgagg tgcagggggc tggcaacgaa aatgagcctc
      661 gcgaggccga caagagccac ccggagcagc gcgagcttcg gcctcggctc tgtaccatga
      721 agaagggccc cgccggctat ggcttcaacc tgcacagcga caagtccaag ccaggccagt
      781 tcatccggtc agtggaccca gactccccgg ctgaggcttc agggctccgg gcccaggatc
      841 gcattgtgga ggtgaacggg gtctgcatgg aggggaagca gcatggggac gtggtgtccg
      901 ccatcagggc tggcggggac gagaccaagc tgctggtggt ggacagggaa actgacgagt
      961 tcttcaagaa atgcagagtg atcccatctc aggagcacct gaatggtccc ctgcctgtgc
     1021 ccttcaccaa tggggagata cagaaggaga acagtcgtga agccctggca gaggcagcct
     1081 tggagagccc caggccagcc ctggtgagat ccgcctccag tgacaccagc gaggagctga
     1141 attcccaaga cagcccccca aaacaggact ccacagcgcc ctcgtctacc tcctcctccg
     1201 accccatcct agacttcaac atctccctgg ccatggccaa agagagggcc caccagaaac
     1261 gcgccgccaa acgggccccg cagatggact ggagcaagaa aaacgaactc ttcagcaacc
     1321 tctgagccgg gtcgagggga attctgcagt cgacggtacc gcgggcccgg gatccaccgg
     1381 atctagataa ctgatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     1441 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     1501 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     1561 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     1621 acgcgtaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     1681 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     1741 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     1801 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     1861 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     1921 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     1981 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2041 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcaggtggc acttttcggg
     2101 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2161 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtcctgagg
     2221 cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc
     2281 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc
     2341 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat
     2401 agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc
     2461 gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga
     2521 gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca aagatcgatc
     2581 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     2641 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     2701 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     2761 acctgtccgg tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca
     2821 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     2881 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     2941 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     3001 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     3061 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     3121 ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     3181 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     3241 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     3301 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     3361 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     3421 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     3481 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     3541 cggggatctc atgctggagt tcttcgccca ccctaggggg aggctaactg aaacacggaa
     3601 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     3661 cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc cagggctggc actctgtcga
     3721 taccccaccg agaccccatt ggggccaata cgcccgcgtt tcttcctttt ccccacccca
     3781 ccccccaagt tcgggtgaag gcccagggct cgcagccaac gtcggggcgg caggccctgc
     3841 catagcctca ggttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     3901 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     3961 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     4021 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     4081 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     4141 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     4201 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     4261 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     4321 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     4381 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     4441 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     4501 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     4561 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     4621 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     4681 ttctgtggat aaccgtatta ccgccatgca ttagttatta atagtaatca attacggggt
     4741 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
     4801 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
     4861 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
     4921 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
     4981 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
     5041 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
     5101 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
     5161 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
     5221 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg
     5281 gtttagtgaa ccgtcagatc cgctagcgct accggtcgcc accatggtga gcaagggcga
     5341 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     5401 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     5461 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     5521 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     5581 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     5641 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     5701 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     5761 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca agg
//
