LOCUS       sequence-109662-        6748 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a Fluorescent Reporter for Peroxynitrite in Mammalian Cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6748)
  AUTHORS   Chen ZJ, Ren W, Wright QE, Ai HW
  TITLE     Genetically encoded fluorescent probe for the selective detection of
            peroxynitrite.
  JOURNAL   J Am Chem Soc. 2013 Oct 9;135(40):14940-3. doi: 10.1021/ja408011q. 
            Epub 2013 Sep 26.
  PUBMED    24059533
REFERENCE   2  (bases 1 to 6748)
  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/ja408011q";
            journalName: "J Am Chem Soc"; date: "2013-10-9- 9"; volume: "135"; 
            issue: "40"; pages: "14940-3"
COMMENT     Sequence Label: pcDNA3-pnGFP
FEATURES             Location/Qualifiers
     source          1..6748
                     /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      898..907
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1137..1158
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1270..1291)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             1663..1680
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    1696..2284
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1749..1769)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2167..2178)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        complement(2300..2318)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2301..2318)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2338..2355)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2344..2568
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2614..3042
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2701..2720)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2911..2932
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3051..3071)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3056..3386
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3237..3372
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3299..3318
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3453..4247
                     /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(3507..3526)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4117..4136
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4421..4542
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4458..4477)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4512..4531
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4591..4607)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4591..4607)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4604..4626)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4615..4631
                     /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(4639..4645,4646..4663,4664..4669))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4684..4705
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4822..4839)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4993..5581)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5073..5092)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5752..6543,6544..6612))
                     /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     6375..6394
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6613..6717)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct 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 atgggtggac 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 gcttgccgcc
      901 accatgggct ccagcactta caacagccac aaggtctata tcaccgccga caagcagaag
      961 aacggcatca aggtgaactt caagatccgc cacaacgtgg aggacggcag cgtgcagctc
     1021 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1081 cactacctga gcacccagtc cgtgctgagc aaagacccca acgagaagcg cgatcacatg
     1141 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1201 gtggacggcg gatccggtgg caccggagtg agcaagggcg aggagctgtt caccggggtg
     1261 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgaggggc
     1321 gaaggcgagg gcgatgccac caacggcaag ctgaccctga agttcatctg caccaccggc
     1381 aagctgcccg tgccctggcc caccctcgtg accaccttga cctagggcgt gcagtgcttc
     1441 tcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     1501 tacgtccagg agcgcaccat cttcttcaag gacgacggca cctacaagac ccgcgccgag
     1561 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     1621 gaggacggca acatcctggg gcacaagctg gagtacaaca cgcaccatca ccatcaccat
     1681 taactcgagg tcgacaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     1741 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     1801 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     1861 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     1921 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     1981 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     2041 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     2101 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     2161 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     2221 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     2281 ccgctctaga gggccctatt ctatagtgtc acctaaatgc tagagctcgc tgatcagcct
     2341 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     2401 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     2461 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     2521 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     2581 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     2641 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     2701 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     2761 taaatcgggg catcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     2821 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     2881 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     2941 tcaaccctat ctcggtctat tcttttgatt tataagggat tttggggatt tcggcctatt
     3001 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
     3061 tcagttaggg tgtggaaagt ccccaggctc cccaggcagg cagaagtatg caaagcatgc
     3121 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3181 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3241 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3301 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3361 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     3421 ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     3481 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     3541 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     3601 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     3661 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     3721 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3781 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3841 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3901 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     3961 tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg
     4021 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     4081 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     4141 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     4201 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     4261 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     4321 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     4381 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     4441 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4501 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     4561 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     4621 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4681 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4741 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4801 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4861 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4921 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4981 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5041 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5101 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5161 cccttcggga agcgtggcgc tttctcaatg ctcacgctgt aggtatctca gttcggtgta
     5221 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5281 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5341 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5401 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     5461 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5521 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5581 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5641 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5701 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5761 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5821 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5881 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5941 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6001 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6061 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6121 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6181 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6241 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6301 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6361 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6421 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6481 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6541 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6601 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6661 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6721 atttccccga aaagtgccac ctgacgtc
//
