LOCUS       sequence-399278-        4773 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses fluorescent protein-tagged lipid biosensor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4773)
  AUTHORS   Wills RC, Doyle CP, Zewe JP, Pacheco J, Hansen SD, Hammond GRV
  TITLE     A novel homeostatic mechanism tunes PI(4,5)P2-dependent signaling at
            the plasma membrane.
  JOURNAL   J Cell Sci. 2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494.
  PUBMED    37534432
REFERENCE   2  (bases 1 to 4773)
  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: "J Cell Sci.
            2023 Aug 3:jcs.261494. doi: 10.1242/jcs.261494."
COMMENT     Sequence Label: PH-PLCd1-mNGx1
FEATURES             Location/Qualifiers
     source          1..4773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        97..196
                     /label=CMVd1 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields slightly reduced expression 
                     relative to the full-length promoter"
     primer_bind     147..167
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             790..1500
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      1045..1054
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    1623..1744
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1660..1679)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1714..1733
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1751..2206)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1888..1909)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2100..2119
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2233..2337
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2339..2696
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2339..2530
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2362..2382)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2547..2682
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2731..3525
                     /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             2731..3525
                     /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(2785..2804)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3395..3414
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3713..3732)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3757..3804
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4133..4721
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4622..4641
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc
       61 ataccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      121 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
      181 ctatataagc agagctggtt tagtgaaccg tcagatccgc tagcgctacc ggactcagat
      241 ctcgagctca agcttcgaat tcacggcatg gactcgggcc gggacttcct gaccctgcac
      301 ggcctacagg atgatgagga tctacaggcg ctgctgaagg gcagccagct cctgaaggtg
      361 aagtccagct catggaggag agagcgcttc tacaagttgc aggaggactg caagaccatc
      421 tggcaggagt cccgcaaggt catgcggacc ccggagtccc agctgttctc catcgaggac
      481 attcaggagg tgcgaatggg gcaccgcacg gagggtctgg agaagttcgc ccgtgatgtg
      541 cccgaggacc gctgcttctc cattgtcttc aaggaccagc gcaatacact agacctcatc
      601 gccccatcgc cagctgatgc ccagcactgg gtgctggggc tgcacaagat catccaccac
      661 tcaggctcca tggaccagcg tcagaagcta cagcactgga ttcactcctg cttgcgaaaa
      721 gctgacaaaa acaaggacaa caagatgagc ttcaaggagc tgcagaactt cctgaagggg
      781 gtgggtggca tggtgagcaa gggcgaggag gataacatgg cctctctccc agcgacacat
      841 gagttacaca tctttggctc catcaacggt gtggactttg acatggtggg tcagggcacc
      901 ggcaatccaa atgatggtta tgaggagtta aacctgaagt ccaccaaggg tgacctccag
      961 ttctccccct ggattctggt ccctcatatc gggtatggct tccatcagta cctgccctac
     1021 cctgacggga tgtcgccttt ccaggccgcc atggtagatg gctccggata ccaagtccat
     1081 cgcacaatgc agtttgaaga tggtgcctcc cttactgtta actaccgcta cacctacgag
     1141 ggaagccaca tcaaaggaga ggcccaggtg aaggggactg gtttccctgc tgacggtcct
     1201 gtgatgacca actcgctgac cgctgcggac tggtgcaggt cgaagaagac ttaccccaac
     1261 gacaaaacca tcatcagtac ctttaagtgg agttacacca ctggaaatgg caagcgctac
     1321 cggagcactg cgcggaccac ctacaccttt gccaagccaa tggcggctaa ctatctgaag
     1381 aaccagccga tgtacgtgtt ccgtaagacg gagctcaagc actccaagac cgagctcaac
     1441 ttcaaggagt ggcaaaaggc ctttaccgat gtgatgggca tggacgagct gtacaagtaa
     1501 agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
     1561 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     1621 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     1681 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     1741 cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
     1801 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     1861 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     1921 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     1981 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     2041 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     2101 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     2161 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
     2221 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     2281 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct
     2341 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     2401 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     2461 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2521 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     2581 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     2641 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc
     2701 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     2761 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     2821 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     2881 accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg
     2941 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     3001 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     3061 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     3121 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     3181 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     3241 ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     3301 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     3361 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     3421 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     3481 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     3541 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     3601 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     3661 agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac
     3721 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     3781 cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
     3841 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
     3901 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
     3961 ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat
     4021 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     4081 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     4141 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     4201 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4261 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4321 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4381 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4441 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4501 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4561 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4621 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4681 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4741 ttttacggtt cctggccttt tgctggcctt ttg
//
