LOCUS       sequence-207862-        5067 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 5067)
  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 5067)
  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-mCherry-P4Mx1
FEATURES             Location/Qualifiers
     source          1..5067
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        49..352
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        353..556
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     507..527
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      583..592
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      640..649
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             646..1353
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(791..809)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1059..1078
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1369..1674
                     /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    1843..1964
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1880..1899)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1934..1953
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1971..2426)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2108..2129)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2320..2339
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2453..2557
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2559..2916
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2559..2750
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2582..2602)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2767..2902
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2951..3745
                     /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             2951..3745
                     /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(3005..3024)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3615..3634
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3933..3952)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3977..4024
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4353..4941
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4842..4861
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
       61 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      121 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      181 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
      241 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
      301 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      361 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      421 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      481 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
      541 ctatataagc agagctggtt tagtgaaccg tcagatccgc tagccaccat ggctctgcag
      601 aaaaagttgg aagagcttga gctggatgag gcaccggtcg ccaccatggt gagcaagggc
      661 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
      721 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
      781 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
      841 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc
      901 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
      961 gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac
     1021 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
     1081 atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag
     1141 atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc
     1201 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac
     1261 atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
     1321 cactccaccg gcggcatgga cgagctgtac aagtccggac tcagatccac ggcaagcacg
     1381 gaaaacttta aaaatgttaa agaaaaatat cagcaaatgc gaggtgatgc tttaaaaaca
     1441 gaaatcctgg ctgatttcaa ggataaactg gctgaagcta ccgatgaaca gagccttaag
     1501 caaattgttg ctgaattgaa aagtaaagat gaatatagaa ttttggctaa ggggcaaggg
     1561 ttaacaaccc agcttttagg gttaaagaca agttcagtgt cttcatttga gaaaatggtt
     1621 gaggaaacaa gagaaagtat taagtctcaa gaaagacaaa ccattaagat aaaataagaa
     1681 ttcccgggtc gactcgacgg taccgcgggc ccgggatcca ccggatctag ataactgatc
     1741 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     1801 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     1861 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     1921 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaacgcgt aaattgtaag
     1981 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     2041 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     2101 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     2161 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     2221 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     2281 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     2341 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     2401 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     2461 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     2521 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     2581 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     2641 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     2701 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     2761 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     2821 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     2881 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     2941 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     3001 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     3061 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     3121 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     3181 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3241 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3301 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3361 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3421 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     3481 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3541 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3601 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3661 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     3721 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     3781 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     3841 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     3901 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     3961 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     4021 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     4081 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     4141 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     4201 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     4261 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     4321 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     4381 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     4441 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     4501 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     4561 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     4621 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     4681 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     4741 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     4801 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     4861 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     4921 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     4981 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     5041 attaccgcca tgcattagtt attaata
//
