LOCUS       sequence-254653-        5299 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of AcrIIC3Nme fused to mCherry with 3x miR-122 
            binding sites in 3' UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5299)
  AUTHORS   Lee J, Mou H, Ibraheim R, Liang SQ, Liu P, Xue W, Sontheimer EJ
  TITLE     Tissue-restricted Genome Editing in vivo Specified by 
            MicroRNA-repressible Anti-CRISPR Proteins.
  JOURNAL   RNA. 2019 Aug 22. pii: rna.071704.119. doi: 10.1261/rna.071704.119.
  PUBMED    31439808
REFERENCE   2  (bases 1 to 5299)
  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: "RNA. 2019 
            Aug 22. pii: rna.071704.119. doi: 10.1261/rna.071704.119."
COMMENT     Sequence Label: p1147-pCSDest-AcrIIC3Nme-mCherry-miR122BS
FEATURES             Location/Qualifiers
     source          1..5299
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        4..22
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     4..21
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             414..437
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             444..464
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             477..1187
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(622..640)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     890..909
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     promoter        complement(1297..1315)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    1320..1454
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1344..1363)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1398..1417
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1567..1587)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1567..1585)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1606..1622)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1606..1622)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1619..1641)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1630..1646
                     /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(1654..1660,1661..1678,1679..1684))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1699..1720
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1837..1854)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2008..2596)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2088..2107)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2767..3558,3559..3627))
                     /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     3390..3409
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3628..3732)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3758..4213)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3895..3916)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4107..4126
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4284..5268
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 ttgatttagg tgacactata gacggaattg tacccgcggc cgatccaccg gtcgccacca
       61 tgttcaaacg ggccattatt ttcaccagct tcaacggctt cgagaaggtc agtcgcacgg
      121 aaaaacggcg ccttgccaag ataattaacg ccagagttag tataatcgac gagtatcttc
      181 gcgccaaaga caccaacgcc tccttggacg gacagtatag agcattcctt ttcaatgatg
      241 agtcaccagc gatgaccgaa ttcctcgcta agctcaaggc gttcgcagaa agctgtactg
      301 gtataagcat tgatgcgtgg gagattgaag agagcgagta tgttcgattg cccgtcgaac
      361 ggagagattt ccttgcggcg gccaatggca aagaaatttt caaaattggc ggggattata
      421 aggatgatga tgataagggc tcccctaaga agaagagaaa ggtgggatcc ggaagtatgg
      481 tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc
      541 acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
      601 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg
      661 cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg
      721 ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga
      781 tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctcactg caggacggcg
      841 agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc
      901 agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc
      961 tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg
     1021 tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca
     1081 tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg
     1141 ctgagggccg ccactccacc ggcggcatgg acgagctgta caagtgataa agcggccatc
     1201 aagcttgaca aacaccattg tcacactcca acaaacacca ttgtcacact ccaacaaaca
     1261 ccattgtcac actccaccat agagctcgct gatcagccta tagtgagtcg tattacgtag
     1321 atccagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     1381 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     1441 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg
     1501 tgtgggaggt tttttaattc gcggccgcgg cgccaatgca ttgggcccgg tacccagctt
     1561 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     1621 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggga gcataaagtg
     1681 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     1741 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     1801 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     1861 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     1921 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     1981 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2041 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2101 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     2161 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2221 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2281 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2341 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2401 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     2461 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     2521 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     2581 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     2641 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     2701 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     2761 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2821 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     2881 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     2941 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3001 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3061 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3121 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     3181 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     3241 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     3301 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     3361 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     3421 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     3481 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     3541 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     3601 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     3661 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     3721 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     3781 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     3841 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     3901 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     3961 tatcagggcg atggcccact acgtgaacca tcaccctaat caaatttttt ggggtcgagg
     4021 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     4081 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     4141 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     4201 ctacagggcg cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     4261 cgggcctctt cgctattacg ccagtcgacc gccaattcaa tatggcgtat atggactcat
     4321 gccaattcaa tatggtggat ctggacctgt gccaattcaa tatggcgtat atggactcgt
     4381 gccaattcaa tatggtggat ctggacccca gccaattcaa tatggcggac ttggcaccat
     4441 gccaattcaa tatggcggac ctggcactgt gccaactggg gaggggtcta cttggcacgg
     4501 tgccaagttt gaggaggggt cttggccctg tgccaagtcc gccatattga attggcatgg
     4561 tgccaataat ggcggccata ttggctatat gccaggatca atatataggc aatatccaat
     4621 atggccctat gccaatatgg ctattggcca ggttcaatac tatgtattgg ccctatgcca
     4681 tatagtattc catatatggg ttttcctatt gacgtagata gcccctccca atgggcggtc
     4741 ccatatacca tatatggggc ttcctaatac cgcccatagc cactccccca ttgacgtcaa
     4801 tggtctctat atatggtctt tcctattgac gtcatatggg cggtcctatt gacgtatatg
     4861 gcgcctcccc cattgacgtc aattacggta aatggcccgc ctggctcaat gcccattgac
     4921 gtcaatagga ccacccacca ttgacgtcaa tgggatggct cattgcccat tcatatccgt
     4981 tctcacgccc cctattgacg tcaatgacgg taaatggccc acttggcagt acatcaatat
     5041 ctattaatag taacttggca agtacattac tattggaagt acgccagggt acattggcag
     5101 tactcccatt gacgtcaatg gcggtaaatg gcccgcgatg gctgccaagt acatccccat
     5161 tgacgtcaat ggggaggggc aatgacgcaa atgggcgttc cattgacgta aatgggcggt
     5221 aggcgtgcct aatgggaggt ctatataagc aatgctcgtt tagggaaccg ccattctgcc
     5281 tggggacgtc ggagcaagc
//
