LOCUS       sequence-364215-        7354 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Genetically encoded FRET-based sensor for monitoring plasma membrane
            PI(4,5)P2 dynamics in living cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7354)
  AUTHORS   Hertel F, Li S, Chen M, Pott L, Mehta S, Zhang J
  TITLE     Fluorescent Biosensors for Multiplexed Imaging of Phosphoinositide 
            Dynamics.
  JOURNAL   ACS Chem Biol. 2020 Jan 17;15(1):33-38. doi: 
            10.1021/acschembio.9b00691. Epub 2020 Jan 6.
  PUBMED    31855412
REFERENCE   2  (bases 1 to 7354)
  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"; doi: 
            "10.1021/acschembio.9b00691"; journalName: "ACS Chem Biol"; date: 
            "2020-01-17- 17"; volume: "15"; issue: "1"; pages: "33-38"
COMMENT     Sequence Label: pcDNA3-Lyn-PlcR
FEATURES             Location/Qualifiers
     source          1..7354
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      901..910
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(946..948,949..951,952..1662)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2194..2196,2197..2199,2200..2913)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2945..2962)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2951..3175
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3221..3649
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3308..3327)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3518..3539
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3658..3678)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3663..3992
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3843..3978
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3905..3924
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4059..4853
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4113..4132)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4723..4742
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5027..5148
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5064..5083)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5118..5137
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5197..5213)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5197..5213)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5210..5232)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5221..5237
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5245..5251,5252..5269,5270..5275))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5290..5311
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5428..5445)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5599..6187)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5679..5698)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6358..7149,7150..7218))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6981..7000
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7219..7323)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gccaccatgg gctgcatcaa gagcaagcgc aaggacaaga agcttatggt gagcaagggc
      961 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     1021 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     1081 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccttc
     1141 ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc
     1201 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     1261 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     1321 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     1381 tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     1441 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     1501 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagctaccag
     1561 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     1621 accgccgccg ggatcactct cggcatggac gagctgtaca agggatccat ggactcgggt
     1681 agggacttcc tgaccctgca cgggctccag gatgacccgg accttcaggc ccttctgaag
     1741 ggcagccagc ttctgaaggt gaagtccagc tcgtggcgta gggaacgctt ctacaagcta
     1801 caggaggact gcaagaccat ctggcaggaa tctcgaaagg tcatgaggtc cccggagtcg
     1861 cagctgttct ccatcgagga cattcaggag gtacggatgg gacaccgcac agaaggcctg
     1921 gagaagtttg cccgagacat ccccgaggat cgatgcttct ccattgtctt caaggaccag
     1981 cgcaacaccc tagacctcat tgccccatca ccagctgacg ctcagcactg ggtgcagggc
     2041 ctgcgcaaga tcatccacca ctccggctcc atggaccagc ggcagaagct gcagcactgg
     2101 attcactcct gcttgcgaaa ggctgatgaa ttcggcggta gcgtggcaga agaggaagat
     2161 gacgaggaag aggacgagga cgatggcggc agcatggtga gcaagggcga ggagctgttc
     2221 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
     2281 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
     2341 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctggggcgtg
     2401 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     2461 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     2521 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     2581 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta catcagccac
     2641 aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc
     2701 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     2761 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     2821 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     2881 atcactctcg gcatggacga gctgtacaag taatctagag ggcccgttta aacccgctga
     2941 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     3001 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     3061 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     3121 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     3181 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     3241 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3301 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3361 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3421 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3481 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3541 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3601 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     3661 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3721 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     3781 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     3841 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     3901 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     3961 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     4021 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     4081 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     4141 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     4201 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     4261 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     4321 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     4381 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     4441 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     4501 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     4561 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     4621 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     4681 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     4741 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     4801 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     4861 tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc
     4921 caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat
     4981 gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc
     5041 agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt
     5101 ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat
     5161 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     5221 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     5281 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     5341 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5401 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5461 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5521 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5581 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5641 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     5701 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     5761 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     5821 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     5881 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5941 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     6001 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     6061 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6121 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6181 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6241 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6301 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6361 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6421 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6481 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6541 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6601 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6661 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     6721 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     6781 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6841 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6901 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6961 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     7021 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     7081 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7141 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7201 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7261 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7321 gcgcacattt ccccgaaaag tgccacctga cgtc
//
