LOCUS       sequence-203961-        5082 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PI4P biosensor .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5082)
  AUTHORS   Zewe JP, Wills RC, Sangappa S, Goulden BD, Hammond GR
  TITLE     SAC1 degrades its lipid substrate PtdIns4P in the endoplasmic 
            reticulum to maintain a steep chemical gradient with donor 
            membranes.
  JOURNAL   Elife. 2018 Feb 20;7. pii: 35588. doi: 10.7554/eLife.35588.
  PUBMED    29461204
REFERENCE   2  (bases 1 to 5082)
  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: "Elife."; 
            date: "2018-02-20"; volume: "7. pii: 35588. doi"; pages: " 
            10.7554/eLife.35588"
COMMENT     Sequence Label: NES-mTagBFP2-P4Mx1
FEATURES             Location/Qualifiers
     source          1..5082
                     /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      595..604
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      652..661
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             658..1359
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIK
                     VVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMAL
                     KLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARY
                     CDLPSKLGHKLN"
     CDS             1432..1737
                     /codon_start=1
                     /product="high-affinity phosphatidylinositol 4-phosphate 
                     binding module of SidM (DrrA) from Legionella pneumophila 
                     (Hammond et al., 2014)"
                     /label=P4M
                     /translation="TASTENFKNVKEKYQQMRGDALKTEILADFKDKLAEATDEQSLKQ
                     IVAELKSKDEYRILAKGQGLTTQLLGLKTSSVSSFEKMVEETRESIKSQERQTIKIK"
     polyA_signal    1870..1991
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1907..1926)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1961..1980
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1998..2453)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2135..2156)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2347..2366
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2480..2584
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2586..2943
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2586..2777
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2609..2629)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2794..2929
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2978..3772
                     /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             2978..3772
                     /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(3032..3051)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3642..3661
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3960..3979)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4004..4051
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4380..4968
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4869..4888
                     /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 gctagccacc
      601 atggctctgc agaaaaagtt ggaagagctt gagctggatg aggcaccggt cgccaccatg
      661 gtgagcgagc tgattaagga gaacatgcac atgaagctgt acatggaggg caccgtggac
      721 aaccatcact tcaagtgcac atccgagggc gaaggcaagc cctacgaggg cacccagacc
      781 atgagaatca aggtggtcga gggcggccct ctccccttcg ccttcgacat cctggctact
      841 agcttcctct acggcagcaa gaccttcatc aaccacaccc agggcatccc cgacttcttc
      901 aagcagtcct tccctgaggg cttcacatgg gagagagtca ccacatacga agacgggggc
      961 gtgctgaccg ctacccagga caccagcctc caggacggct gcctcatcta caacgtcaag
     1021 atcagagggg tgaacttcac atccaacggc cctgtgatgc agaagaaaac actcggctgg
     1081 gaggccttca ccgagacgct gtaccccgct gacggcggcc tggaaggcag aaacgacatg
     1141 gccctgaagc tcgtgggcgg gagccatctg atcgcaaacg ccaagaccac atatagatcc
     1201 aagaaacccg ctaagaacct caagatgcct ggcgtctact atgtggacta cagactggaa
     1261 agaatcaagg aggccaacaa cgagacctac gtcgagcagc acgaggtggc agtggccaga
     1321 tactgcgacc tccctagcaa actggggcac aagcttaatt ccggactcag atctcgagct
     1381 caagcttcga attctgcagt cgacggtggt agtgcttccg gactcagatc cacggcaagc
     1441 acggaaaact ttaaaaatgt taaagaaaaa tatcagcaaa tgcgaggtga tgctttaaaa
     1501 acagaaatcc tggctgattt caaggataaa ctggctgaag ctaccgatga acagagcctt
     1561 aagcaaattg ttgctgaatt gaaaagtaaa gatgaatata gaattttggc taaggggcaa
     1621 gggttaacaa cccagctttt agggttaaag acaagttcag tgtcttcatt tgagaaaatg
     1681 gttgaggaaa caagagaaag tattaagtct caagaaagac aaaccattaa gataaaataa
     1741 ggatccaccg gatctagata actgatcata atcagccata ccacatttgt agaggtttta
     1801 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     1861 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     1921 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     1981 aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     2041 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     2101 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     2161 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     2221 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     2281 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     2341 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     2401 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg
     2461 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2521 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     2581 gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     2641 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     2701 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     2761 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     2821 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     2881 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     2941 aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     3001 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     3061 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     3121 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc
     3181 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     3241 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     3301 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     3361 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     3421 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     3481 cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca
     3541 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     3601 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     3661 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     3721 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     3781 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     3841 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     3901 atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact
     3961 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     4021 aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     4081 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     4141 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     4201 gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat
     4261 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     4321 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4381 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4441 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4501 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4561 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4621 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4681 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4741 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4801 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4861 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4921 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4981 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     5041 ttatcccctg attctgtgga taaccgtatt accgccatgc at
//
