LOCUS       sequence-329343-        6747 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses two tandem repeats of Tapp1 PH domain that binds to 
            PI(3,4)P2. Fused to eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6747)
  AUTHORS   Roman-Fernandez A, Roignot J, Sandilands E, Nacke M, Mansour MA, 
            McGarry L, Shanks E, Mostov KE, Bryant DM
  TITLE     The phospholipid PI(3,4)P2 is an apical identity determinant.
  JOURNAL   Nat Commun. 2018 Nov 28;9(1):5041. doi: 10.1038/s41467-018-07464-8.
  PUBMED    30487552
REFERENCE   2  (bases 1 to 6747)
  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"; journalName: "Nat 
            Commun."; date: "2018-11-28"; volume: "9(1):5041. doi"; pages: " 
            10.1038/s41467-018-07464-8"
COMMENT     Sequence Label: GFP-2xPH-TAPP1 WT
FEATURES             Location/Qualifiers
     source          1..6747
                     /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"
     regulatory      2302..2311
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2311..2340
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    3535..3656
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3572..3591)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3626..3645
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3663..4118)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3800..3821)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4012..4031
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4145..4249
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4251..4608
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4251..4442
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4274..4294)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4459..4594
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4643..5437
                     /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             4643..5437
                     /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(4697..4716)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5307..5326
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5625..5644)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5669..5716
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6045..6633
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6534..6553
                     /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 atctcgagtg aatgatcagc aggacctagt ggaatgggta
     1381 aatgtgttaa acaaagctat aaaaattaca gtaccaaagc agtcagactc acagcctaat
     1441 tctgataacc taagtcgcca tggtgaatgt gggaaaaagc aagtgtctta cagaactgat
     1501 attgttggtg gcgtacccat cattactccc actcagaaag aagaagtaaa tgaatgtggt
     1561 gaaagtattg acagaaataa tctgaaacgg tcacaaagcc atcttcctta ctttactcct
     1621 aaaccacctc aagatagtgc ggttatcaaa gctggatatt gtgtaaaaca aggagcagtg
     1681 atgaaaaact ggaagagaag atattttcaa ttggatgaaa acacaatagg ctacttcaaa
     1741 tctgaactgg aaaaggaacc tcttcgcgta ataccactta aagaggttca taaagtccag
     1801 gaatgtaagc aaagcgacat aatgatgagg gacaacctct ttgaaattgt aacaacgtct
     1861 cgaactttct atgtgcaggc tgatagccct gaagagatgc acagttggat taaagcagtc
     1921 tctggcgcca ttgtagcaca gcggggtccc ggcagatctg cgtcttctga gcatcccccc
     1981 ggtccttcag aatccaaaca cgctttccgt cctaccaacg cagccgccgc cacctcacat
     2041 tccacagcct ctcgcagcaa ctctttggtc tcaaccttta ccatggagaa gcgaggattt
     2101 tacgagtctc ttgccaaggt caagccaggg aacttcaagg tccagactgt ctctccaaga
     2161 gaaccagctt ccaaagtgac tgaacaagct ctgttaagac ctcaaagtaa aaatggccct
     2221 caggaaaaag attgtgacct agtagacttg gacgatgcga gccttccggt cagtgacgtg
     2281 ggtaccgcgg gcccgggatc cgccaccatg gaacaaaaac tcatctcaga agaggatctg
     2341 aatgatcagc aggacctagt ggaatgggta aatgtgttaa acaaagctat aaaaattaca
     2401 gtaccaaagc agtcagactc acagcctaat tctgataacc taagtcgcca tggtgaatgt
     2461 gggaaaaagc aagtgtctta cagaactgat attgttggtg gcgtacccat cattactccc
     2521 actcagaaag aagaagtaaa tgaatgtggt gaaagtattg acagaaataa tctgaaacgg
     2581 tcacaaagcc atcttcctta ctttactcct aaaccacctc aagatagtgc ggttatcaaa
     2641 gctggatatt gtgtaaaaca aggagcagtg atgaaaaact ggaagagaag atattttcaa
     2701 ttggatgaaa acacaatagg ctacttcaaa tctgaactgg aaaaggaacc tcttcgcgta
     2761 ataccactta aagaggttca taaagtccag gaatgtaagc aaagcgacat aatgatgagg
     2821 gacaacctct ttgaaattgt aacaacgtct cgaactttct atgtgcaggc tgatagccct
     2881 gaagagatgc acagttggat taaagcagtc tctggcgcca ttgtagcaca gcggggtccc
     2941 ggcagatctg cgtcttctga gcatcccccc ggtccttcag aatccaaaca cgctttccgt
     3001 cctaccaacg cagccgccgc cacctcacat tccacagcct ctcgcagcaa ctctttggtc
     3061 tcaaccttta ccatggagaa gcgaggattt tacgagtctc ttgccaaggt caagccaggg
     3121 aacttcaagg tccagactgt ctctccaaga gaaccagctt ccaaagtgac tgaacaagct
     3181 ctgttaagac ctcaaagtaa aaatggccct caggaaaaag attgtgacct agtagacttg
     3241 gacgatgcga gccttccggt cagtgacgtg tgaggcagaa gcgcacggag cctgcctgcc
     3301 tctgccgtcc tcagttacct ttcatgaggc ttctagccaa agatgataaa gggggaaatg
     3361 gtttttagtg cgtatattat actgcctctt aggtgtactc tttgaggatc caccggatct
     3421 agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     3481 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     3541 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3601 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc
     3661 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     3721 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     3781 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     3841 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     3901 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     3961 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     4021 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     4081 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     4141 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4201 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     4261 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     4321 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     4381 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     4441 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     4501 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     4561 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     4621 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     4681 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     4741 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     4801 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     4861 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     4921 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4981 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     5041 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     5101 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     5161 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     5221 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     5281 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     5341 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     5401 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     5461 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     5521 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     5581 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     5641 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     5701 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     5761 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     5821 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     5881 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5941 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     6001 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6061 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6121 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     6181 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     6241 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6301 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     6361 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     6421 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6481 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6541 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6601 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     6661 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     6721 gtggataacc gtattaccgc catgcat
//
