LOCUS       sequence-249192-        4925 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses AcrIIC1(N3F/D15Q/A48I) fused to mCherry; mediates potent 
            inhibition of both, N. meningitidis as well as S. aureus Cas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4925)
  AUTHORS   Mathony J, Harteveld Z, Schmelas C, Upmeier Zu Belzen J, 
            Aschenbrenner S, Sun W, Hoffmann MD, Stengl C, Scheck A, Georgeon S,
            Rosset S, Wang Y, Grimm D, Eils R, Correia BE, Niopek D
  TITLE     Computational design of anti-CRISPR proteins with improved 
            inhibition potency.
  JOURNAL   Nat Chem Biol. 2020 Apr 13. pii: 10.1038/s41589-020-0518-9. doi: 
            10.1038/s41589-020-0518-9.
  PUBMED    32284602
REFERENCE   2  (bases 1 to 4925)
  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: "Nat Chem 
            Biol. 2020 Apr 13. pii: 10.1038/s41589-020-0518-9. doi: 
            10.1038/s41589-020-0518-9."
COMMENT     Sequence Label: AcrIIC1X* (AcrIIC1X-mCherry chimera)
FEATURES             Location/Qualifiers
     source          1..4925
                     /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"
     regulatory      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             822..1526
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(964..982)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1232..1251
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1713..1834
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1750..1769)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1804..1823
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1841..2296)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1978..1999)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2190..2209
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2323..2427
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2429..2786
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2429..2620
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2452..2472)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2637..2772
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2821..3615
                     /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             2821..3615
                     /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(2875..2894)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3485..3504
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3803..3822)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3847..3894
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4223..4811
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4712..4731
                     /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 gctagcgcca
      601 ccatggcatt caaaacgtac aagataggca aaaacgctgg gtaccagggg tgtggtctgt
      661 gcctcgctgc gatcagcgag aatgaggcta tcaaggtcaa atacttgcgc gacatctgcc
      721 cggactacga tggcgacgac aaaatcgaag attggttgag gtgggggacg gattctcgcg
      781 ttaaggccgc tgcgcttgaa atggagcaat atggctccgg cgtgagcaag ggcgaggagg
      841 ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga
      901 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
      961 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc
     1021 ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact
     1081 tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     1141 gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga
     1201 agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct
     1261 gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc
     1321 agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg
     1381 ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct
     1441 cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca
     1501 ccggcggcat ggacgagctg tacaagggct ccggcgccta cacatccgtc ggtatggcgt
     1561 cctgttggga gttcgttgag ttgtgactcg agcggccgcg actctagatc ataatcagcc
     1621 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
     1681 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     1741 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     1801 gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag cgttaatatt
     1861 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     1921 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     1981 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     2041 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     2101 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     2161 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     2221 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     2281 ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2341 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     2401 cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt
     2461 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     2521 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     2581 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     2641 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     2701 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     2761 tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc
     2821 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     2881 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     2941 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     3001 caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     3061 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     3121 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     3181 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     3241 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     3301 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac
     3361 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     3421 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     3481 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     3541 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     3601 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     3661 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     3721 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     3781 cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg
     3841 ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac
     3901 gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc
     3961 aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag
     4021 ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac
     4081 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4141 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4201 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4261 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     4321 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4381 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4441 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4501 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4561 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4621 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4681 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4741 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4801 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4861 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca
     4921 tgcat
//
