LOCUS       sequence-272423-        6215 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CNC3 (CASANOVA-C3), a blue light dependent NmeCas9 
            inhibitory protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6215)
  AUTHORS   Hoffmann MD, Mathony J, Upmeier Zu Belzen J, Harteveld Z, 
            Aschenbrenner S, Stengl C, Grimm D, Correia BE, Eils R, Niopek D
  TITLE     Optogenetic control of Neisseria meningitidis Cas9 genome editing 
            using an engineered, light-switchable anti-CRISPR protein.
  JOURNAL   Nucleic Acids Res. 2021 Mar 18;49(5):e29. doi: 10.1093/nar/gkaa1198.
  PUBMED    33330940
REFERENCE   2  (bases 1 to 6215)
  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: "Nucleic 
            Acids Res."; date: "2021-03-18"; volume: "49(5):e29. doi"; pages: " 
            10.1093/nar/gkaa1198"
COMMENT     Sequence Label: CNC3 (CASANOVA-C3)
FEATURES             Location/Qualifiers
     source          1..6215
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        129..332
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     283..303
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     377..396
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        377..395
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      415..424
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1320..1337)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1326..1550
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1596..2024
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1683..1702)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1893..1914
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2033..2053)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2038..2367
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2218..2353
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2280..2299
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2434..3228
                     /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(2488..2507)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3098..3117
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3402..3523
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3439..3458)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3493..3512
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3572..3588)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3572..3588)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3585..3607)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3596..3612
                     /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(3620..3626,3627..3644,3645..3650))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3665..3686
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3803..3820)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3974..4562)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4054..4073)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4733..5524,5525..5593))
                     /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     5356..5375
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5594..5698)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5773..5792)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(5964..6215,1..128)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
       61 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      121 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
      181 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
      241 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
      301 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt
      361 actggcttat cgaaattaat acgactcact atagggagac ccaagctggc tagcgccacc
      421 atggctttca agagagctat tatatttacg tcctttaatg gttttgagaa ggtctctcga
      481 acagagaagc gacgccttgc caagataatt aacgcgaggg tctcaattat tgatgagtat
      541 ctcagagcca aggatacaaa cgcgagcctg gatggccaat accgagcctt tctgtttctt
      601 gctacaactt tggagcggat tgagaagaat ttcgtcataa ctgatcccag gttgcctgat
      661 aatcctataa tattcgcctc cgatagtttt cttcaactta cggagtactc cagagaggag
      721 atactgggaa ggaattgcag attcctccaa ggtccagaaa ctgatagagc tactgtgagg
      781 aagatacggg atgccattga caatcaaact gaagtgacag tgcaacttat caattacacc
      841 aagtcaggga aaaaattctg gaacctgttc catttgcaac ccatgaggga ccaaaagggc
      901 gatgtgcaat atttcatagg cgttcaactt gatggtactg aacacgtcag agatgcggcg
      961 gagcgagaag gagtgatgct gattaagaaa acagcagaga atatcgacga agcggcgaaa
     1021 gagttgaacg acgaaagccc cgcgatgaca gagttcctcg caaaactgaa agcattcgcg
     1081 gagagctgca cgggaatatc aatagatgcg tgggagattg aagagagcga gtatgttcga
     1141 ctgcccgttg aacgccggga tttcctggcc gctgcgaatg gcaaggaaat cttcaagata
     1201 taactcgagc ggccgccact gtgctggata tctgcagaat tagagacgcc gttgatgttg
     1261 aagtttatat accgttgatg ttgaagtcgt ctctagctta agtttaaacc gctgatcagc
     1321 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     1381 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     1441 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     1501 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     1561 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     1621 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1681 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1741 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     1801 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     1861 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     1921 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     1981 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     2041 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2101 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     2161 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     2221 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     2281 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     2341 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     2401 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     2461 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     2521 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     2581 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     2641 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     2701 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     2761 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     2821 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     2881 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     2941 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     3001 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3061 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3121 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3181 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3241 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3301 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3361 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     3421 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3481 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     3541 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     3601 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3661 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3721 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3781 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3841 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3901 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3961 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4021 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4081 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4141 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4201 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4261 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4321 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4381 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4441 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4501 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4561 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4621 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4681 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4741 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4801 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4861 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4921 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4981 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5041 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5101 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5161 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5221 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5281 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5341 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5401 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5461 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5521 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5581 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5641 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5701 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
     5761 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
     5821 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
     5881 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
     5941 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
     6001 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     6061 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     6121 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     6181 aactgcccac ttggcagtac atcaagtgta tcata
//
