LOCUS       sequence-399528-        5496 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 5496)
  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 5496)
  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-mNGx2
FEATURES             Location/Qualifiers
     source          1..5496
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..134
                     /label=CMVd1 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields slightly reduced expression 
                     relative to the full-length promoter"
     primer_bind     85..105
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             728..1435
                     /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      983..992
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1445..1454
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1451..2161
                     /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      1706..1715
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2284..2405
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2321..2340)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2375..2394
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2412..2867)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2549..2570)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2761..2780
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2894..2998
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3000..3357
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3000..3191
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3023..3043)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3208..3343
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3392..4186
                     /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             3392..4186
                     /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(3446..3465)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4056..4075
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4374..4393)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4418..4465
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4794..5382
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5283..5302
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
       61 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
      121 atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg actcagatct
      181 cgagctcaag cttcgaattc acggcatgga ctcgggccgg gacttcctga ccctgcacgg
      241 cctacaggat gatgaggatc tacaggcgct gctgaagggc agccagctcc tgaaggtgaa
      301 gtccagctca tggaggagag agcgcttcta caagttgcag gaggactgca agaccatctg
      361 gcaggagtcc cgcaaggtca tgcggacccc ggagtcccag ctgttctcca tcgaggacat
      421 tcaggaggtg cgaatggggc accgcacgga gggtctggag aagttcgccc gtgatgtgcc
      481 cgaggaccgc tgcttctcca ttgtcttcaa ggaccagcgc aatacactag acctcatcgc
      541 cccatcgcca gctgatgccc agcactgggt gctggggctg cacaagatca tccaccactc
      601 aggctccatg gaccagcgtc agaagctaca gcactggatt cactcctgct tgcgaaaagc
      661 tgacaaaaac aaggacaaca agatgagctt caaggagctg cagaacttcc tgaagggggt
      721 gggtggcatg gtgagcaagg gcgaggagga taacatggcc tctctcccag cgacacatga
      781 gttacacatc tttggctcca tcaacggtgt ggactttgac atggtgggtc agggcaccgg
      841 caatccaaat gatggttatg aggagttaaa cctgaagtcc accaagggtg acctccagtt
      901 ctccccctgg attctggtcc ctcatatcgg gtatggcttc catcagtacc tgccctaccc
      961 tgacgggatg tcgcctttcc aggccgccat ggtagatggc tccggatacc aagtccatcg
     1021 cacaatgcag tttgaagatg gtgcctccct tactgttaac taccgctaca cctacgaggg
     1081 aagccacatc aaaggagagg cccaggtgaa ggggactggt ttccctgctg acggtcctgt
     1141 gatgaccaac tcgctgaccg ctgcggactg gtgcaggtcg aagaagactt accccaacga
     1201 caaaaccatc atcagtacct ttaagtggag ttacaccact ggaaatggca agcgctaccg
     1261 gagcactgcg cggaccacct acacctttgc caagccaatg gcggctaact atctgaagaa
     1321 ccagccgatg tacgtgttcc gtaagacgga gctcaagcac tccaagaccg agctcaactt
     1381 caaggagtgg caaaaggcct ttaccgatgt gatgggcatg gacgagctgt acaagtcacc
     1441 ggtcgccacc atggtgagca agggcgagga ggataacatg gcctctctcc cagcgacaca
     1501 tgagttacac atctttggct ccatcaacgg tgtggacttt gacatggtgg gtcagggcac
     1561 cggcaatcca aatgatggtt atgaggagtt aaacctgaag tccaccaagg gtgacctcca
     1621 gttctccccc tggattctgg tccctcatat cgggtatggc ttccatcagt acctgcccta
     1681 ccctgacggg atgtcgcctt tccaggccgc catggtagat ggctccggat accaagtcca
     1741 tcgcacaatg cagtttgaag atggtgcctc ccttactgtt aactaccgct acacctacga
     1801 gggaagccac atcaaaggag aggcccaggt gaaggggact ggtttccctg ctgacggtcc
     1861 tgtgatgacc aactcgctga ccgctgcgga ctggtgcagg tcgaagaaga cttaccccaa
     1921 cgacaaaacc atcatcagta cctttaagtg gagttacacc actggaaatg gcaagcgcta
     1981 ccggagcact gcgcggacca cctacacctt tgccaagcca atggcggcta actatctgaa
     2041 gaaccagccg atgtacgtgt tccgtaagac ggagctcaag cactccaaga ccgagctcaa
     2101 cttcaaggag tggcaaaagg cctttaccga tgtgatgggc atggacgagc tgtacaagta
     2161 aagcggccgc gactctagat cataatcagc cataccacat ttgtagaggt tttacttgct
     2221 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     2281 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     2341 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     2401 tcttaaggcg taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     2461 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2521 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2581 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2641 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2701 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2761 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2821 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt
     2881 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     2941 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtcc
     3001 tgaggcggaa agaaccagct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     3061 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     3121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     3241 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     3301 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaagat
     3361 cgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat tgcacgcagg
     3421 ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg
     3481 ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa
     3541 gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc tatcgtggct
     3601 ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga
     3661 ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc
     3721 cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac
     3781 ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc
     3841 cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact
     3901 gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga cccatggcga
     3961 tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg
     4021 ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga
     4081 agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga
     4141 ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg gactctgggg
     4201 ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga ttccaccgcc
     4261 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     4321 cagcgcgggg atctcatgct ggagttcttc gcccacccta gggggaggct aactgaaaca
     4381 cggaaggaga caataccgga aggaacccgc gctatgacgg caataaaaag acagaataaa
     4441 acgcacggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     4501 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     4561 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     4621 cctgccatag cctcaggtta ctcatatata ctttagattg atttaaaact tcatttttaa
     4681 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4741 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4801 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4861 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4921 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     4981 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     5041 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     5101 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     5161 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     5221 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     5281 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     5341 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     5401 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     5461 cctgattctg tggataaccg tattaccgcc atgcat
//
