LOCUS       sequence-69554-e        5896 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Genetically encoded fluorescent indicator for simultaneous PIP3 and 
            hydrogen peroxide detection..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5896)
  AUTHORS   Mishina NM, Bogeski I, Bolotin DA, Hoth M, Niemeyer BA, Schultz C, 
            Zagaynova EV, Lukyanov S, Belousov VV
  TITLE     Can we see PIP(3) and hydrogen peroxide with a single probe?
  JOURNAL   Antioxid Redox Signal. 2012 Aug 1;17(3):505-12. doi: 
            10.1089/ars.2012.4574. Epub 2012 Apr 10.
  PUBMED    22369174
REFERENCE   2  (bases 1 to 5896)
  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: 
            "10.1089/ars.2012.4574"; journalName: "Antioxid Redox Signal"; date:
            "2012-08-1- 1"; volume: "17"; issue: "3"; pages: "505-12"
COMMENT     Sequence Label: pC1-PIP-SHOW
FEATURES             Location/Qualifiers
     source          1..5896
                     /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"
     CDS             1128..2561
                     /codon_start=1
                     /product="improved fluorescent indicator for intracellular 
                     H2O2 (Bilan et al., 2013)"
                     /label=HyPer-3
                     /translation="EMASQQGETMSGPLHIGLIPTVGPYLLPHIIPMLHQTFPKLEMYL
                     HEAQTHQLLAQLDSGKLDCVILALVKESEAFIEVPLFDEPMLLAIYEDHPWANRECVPM
                     ADLAGEKLLMLEDGHCLRDQAMSAGYNSDNVYIMADKQKNGIKANFKIRHNVEDGSVQL
                     ADHYQQNTPIGDGPVLLPDNHYLSFQSVLSKDPNEKRDHMVLLEFVTAAGITLGMDELY
                     NVDGGSGGTGSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKLICTT
                     GKLPVPWPTLVTTLGYGLKCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGNYKTR
                     AEVKFEGDTLVNRIELKGIGFKEDGNILGHKLEYNGTGFCFEAGADEDTHFRATSLETL
                     RNMVAAGSGITLLPALAVPPERKRDGVVYLPCIKPEPRRTIGLVYRPGSPLRSRYEQLA
                     EAIRARMDGHFDKVLKQAV"
     primer_bind     1733..1754
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1863..1884)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2124..2143)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     polyA_signal    2684..2805
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2721..2740)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2775..2794
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2812..3267)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2949..2970)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3161..3180
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3294..3398
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3400..3757
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3400..3591
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3423..3443)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3608..3743
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3792..4586
                     /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             3792..4586
                     /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(3846..3865)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4456..4475
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4774..4793)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4818..4865
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5194..5782
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5683..5702
                     /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 gctagcatgg
      601 ccgcagtgat tctggagagc atctttctga agcgatccca acagaaaaag aaaacatcac
      661 ctctaaactt caagaagcgc ctgtttctct tgaccgtgca caaactctcc tactataagt
      721 atgactttga acgtgggaga agaggcagta agaagggttc aatagatgtt gagaagatca
      781 cttgtgttga aacagtggtt cctgaaaaaa atcctcctcc agaaagacag attccgagaa
      841 gaggtgaaga gtccagcgaa atggagcaaa tttcaatcat tgaaaggttc ccttatccct
      901 tccaggttgt atatgatgaa gggcctctct acgtcttctc cccaactgaa gaactaagga
      961 agcggtggat tcaccagctc aaaaacgtaa tccggtacaa cagtgatctg gttcagaaat
     1021 atcacccttg cttctggatc gatgggcagt atctctgctg ctctcagaca gccaaaaatg
     1081 ctatgggctg ccaaattttg gagaacgggg atccaccggt cgccaccgag atggcaagcc
     1141 agcagggcga gacgatgtcc ggaccgctgc acattggttt gattcccaca gttggaccgt
     1201 acctgctacc gcatattatc cctatgctgc accagacctt tccaaagctg gaaatgtatc
     1261 tgcatgaagc acagacccac cagttactgg cgcaactgga cagcggcaaa ctcgattgcg
     1321 tgatcctcgc gctggtgaaa gagagcgaag cattcattga agtgccgttg tttgatgagc
     1381 caatgttgct ggctatctat gaagatcacc cgtgggcgaa ccgcgaatgc gtaccgatgg
     1441 ccgatctggc aggggaaaaa ctgctgatgc tggaagatgg tcactgtttg cgcgatcagg
     1501 caatgtccgc cggctacaac agcgacaacg tctatatcat ggccgacaag cagaagaacg
     1561 gcatcaaggc caacttcaag atccgccaca acgtcgagga cggcagcgtg cagctcgccg
     1621 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1681 acctgagctt ccagtccgtc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1741 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaacgtgg
     1801 atggcggtag cggtggcacc ggcagcaagg gcgaggagct gttcaccggg gtggtgccca
     1861 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     1921 agggcgatgc cacctacggc aagctgaccc tgaagctgat ctgcaccacc ggcaagctgc
     1981 ccgtgccctg gcccaccctc gtgaccaccc tcggctacgg cctgaagtgc ttcgcccgct
     2041 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     2101 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     2161 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcggcttc aaggaggacg
     2221 gcaacatcct ggggcacaag ctggagtaca acggcaccgg tttctgtttt gaagccgggg
     2281 cggatgaaga tacacacttc cgcgcgacca gcctggaaac tctgcgcaac atggtggcgg
     2341 caggtagcgg gatcacttta ctgccagcgc tggctgtgcc gccggagcgc aaacgcgatg
     2401 gggttgttta tctgccgtgc attaagccgg aaccacgccg cactattggc ctggtttatc
     2461 gtcctggctc accgctgcgc agccgctatg agcagctggc agaggccatc cgcgcaagaa
     2521 tggatggcca tttcgataaa gttttaaaac aggcggttta aagcggccgc gactctagat
     2581 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
     2641 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
     2701 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     2761 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa
     2821 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     2881 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     2941 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3001 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3061 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     3121 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     3181 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     3241 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
     3301 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3361 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
     3421 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3481 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     3541 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     3601 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     3661 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     3721 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     3781 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     3841 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     3901 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     3961 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4021 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4081 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4141 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4201 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4261 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     4321 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4381 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     4441 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     4501 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     4561 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     4621 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     4681 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     4741 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     4801 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     4861 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     4921 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     4981 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     5041 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5101 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5161 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5221 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5281 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5341 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5401 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5461 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5521 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5581 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5641 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5701 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5761 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5821 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     5881 tattaccgcc atgcat
//
