LOCUS       sequence-421874-        5579 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Peroxidase (APEX) reporter for TEV protease cleavage.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5579)
  AUTHORS   Zhou G, Wan WW, Wang W
  TITLE     Modular Peroxidase-Based Reporters for Detecting Protease Activity 
            and Protein Interactions with Temporal Gating.
  JOURNAL   J Am Chem Soc. 2022 Dec 21;144(50):22933-22940. doi: 
            10.1021/jacs.2c08280. Epub 2022 Dec 13.
  PUBMED    36511757
REFERENCE   2  (bases 1 to 5579)
  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/jacs.2c08280"; journalName: "J Am Chem Soc"; date: 
            "2022-12-21- 21"; volume: "144"; issue: "50"; pages: "22933-22940"
COMMENT     Sequence Label: CLAPon
FEATURES             Location/Qualifiers
     source          1..5579
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        201..504
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        505..708
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     659..679
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     753..772
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        753..771
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             896..916
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(1220..1222,1223..1933)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1220..1222,1223..1933)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1262..1283)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1523..1542)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1870..1891
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2600..2617)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2606..2813
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2842..2982
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2842..2971
                     /label=AAV2 ITR
     rep_origin      3057..3512
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3144..3163)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3354..3375
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3529..3548)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3648..3670
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3708..3726)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3794..3898
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3899..3967,3968..4759)
                     /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     complement(4117..4136)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4930..5518
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5419..5438
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgttgtaca tagtaatcaa ttacggggtc attagttcat
      181 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      241 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      301 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      361 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      421 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      481 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      541 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      601 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      661 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      721 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      781 gcttatgatg gaagtcagcg ccttggagaa ggaggtctct gcccttgaaa aagaggtttc
      841 agccctcgag aaagaagtct ctgcgctcga aaaagaagtg tcagcgcttg agaaggagaa
      901 cctgtacttc cagggcaccg gtgggctgct gcagctgccc agcgacaaag ccctgctgtc
      961 cgatcccgtg ttcagacctc tggtcgataa gtatgcagcc gacgaggatg cttttttcgc
     1021 agattacgca gaagcacatc agaagctgtc agaactggga tttgccgacg ccgctagcaa
     1081 agtttccgcc ctcaaggaaa aggtcagtgc tcttaaagag aaagtatccg cactcaaaga
     1141 gaaggttagc gcactgaaag aaaaggtcag cgcactgaaa gaaggatcct ctggttccgg
     1201 aagtggcgga ggagggtcag tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     1261 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     1321 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     1381 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     1441 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     1501 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     1561 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     1621 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     1681 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     1741 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     1801 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     1861 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     1921 cgagctgtac aagtccggtg gctctgggag ctctagtgga tcgactagtg ggaaatcata
     1981 cccaacagtg tccgcagact accaggatgc cgtggagaaa gccaagaaga ggctgggagg
     2041 gtttatcgca gaaaagaggt gcgcacctct gatgctgaga ctggctttcc acagcgcagg
     2101 cacctttgac aagcgaacca aaacaggcgg accctttgga acaatccggc gccctgctga
     2161 actggcacat agtgccaaca gtgggctgga catcgccgtg cggctgctgg aacctctgaa
     2221 agcagagttc ccaattctgt cctacgccga tttttatcag ctggcaggag tggtcgctgt
     2281 ggaggtcact gggggcccca aggtgccttt ccacccagga cgggaggaca aaccagaacc
     2341 acctccagag gggcgcctgc cagatccgac aaagggctcc gaccatctgc gagatgtgtt
     2401 tgggaaagct atgggcctga ctgaccagga tatcgtcgca ctgtctggag ggcacaccct
     2461 tggcgccgct cataaggaaa ggtcaggctt cgagggaccc tggacaagca accccctgat
     2521 tttcgacaat tcttacttta ctgaactgct gagtggagag aaggaataag aattcctcga
     2581 gctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     2641 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     2701 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     2761 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggaggtaacc
     2821 acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     2881 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     2941 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
     3001 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
     3061 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     3121 acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     3181 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3241 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     3301 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3361 cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg
     3421 gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3481 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
     3541 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     3601 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     3661 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     3721 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     3781 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3841 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     3901 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     3961 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     4021 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     4081 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     4141 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     4201 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     4261 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     4321 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     4381 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     4441 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     4501 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     4561 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg
     4621 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     4681 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     4741 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     4801 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     4861 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     4921 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4981 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     5041 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     5101 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     5161 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     5221 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     5281 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     5341 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     5401 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     5461 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     5521 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
