LOCUS       sequence-336857-        5754 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying the MCP-RESCUE-DDN (RESCUE is the evolved ADAR2 
            variant for C-U editing) with a nuclear export signal.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5754)
  AUTHORS   Katrekar D, Xiang Y, Palmer N, Saha A, Meluzzi D, Mali P
  TITLE     Comprehensive interrogation of the ADAR2 deaminase domain for 
            engineering enhanced RNA editing activity and specificity.
  JOURNAL   Elife. 2022 Jan 19;11. pii: 75555. doi: 10.7554/eLife.75555.
  PUBMED    35044296
REFERENCE   2  (bases 1 to 5754)
  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: "Elife."; 
            date: "2022-01-19"; volume: "11. pii: 75555. doi"; pages: " 
            10.7554/eLife.75555"
COMMENT     Sequence Label: Split-RESCUE-N
FEATURES             Location/Qualifiers
     source          1..5754
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     4..20
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   24..164
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   35..164
                     /label=AAV2 ITR
     promoter        183..431
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     183..203
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     354..373
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        433..507
                     /label=Cj gRNA scaffold
                     /note="guide RNA scaffold for the Campylobacter jejuni 
                     CRISPR/Cas9 system (Kim et al., 2016)"
     promoter        592..905
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     824..844
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(917..941)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        956..1335
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1336..1539
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1490..1510
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1536..1560
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1660..1686
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             1693..1716
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2675..2694)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2719..2767
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   2982..3122
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2982..3111
                     /label=AAV2 ITR
     primer_bind     complement(3162..3178)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3162..3178)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3175..3197)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3186..3202
                     /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(3210..3216,3217..3234,3235..3240))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3255..3276
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3564..4152)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3644..3663)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4323..5138)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4323..5138)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5046..5065
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(5139..5243)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5311..5329
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5367..5389)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5489..5508
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5702..5724
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5717..5733
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gttgtaaaac gacggccagt gaacctgcag gcagctgcgc gctcgctcgc tcactgaggc
       61 cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg
      121 agcgcgcaga gagggagtgg ccaactccat cactaggggt tcctgcggcc gcacgcgtgg
      181 aggagggcct atttcccatg attccttcat atttgcatat acgatacaag gctgttagag
      241 agataattag aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta
      301 gaaagtaata atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc
      361 atatgcttac cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag
      421 gacgaaacac cggttttagt ccctgaagga agggactaaa ataaagagtt tgcgggactc
      481 tgcggggtta caatccccta aaaccgctaa ggctagtccg ttatcaactt gaaaaagtgg
      541 caccgagtcg gtgctttttt gctagcgtac tgagtcgccc agtctcagat agatccgacg
      601 ccgccatctc taggcccgcg ccggccccct cgcacagact tgtgggagaa gctcggctac
      661 tcccctgccc cggttaattt gcatataata tttcctagta actatagagg cttaatgtgc
      721 gataaaagac agataatctg ttctttttaa tactagctac attttacatg ataggcttgg
      781 atttctataa gagatacaaa tactaaatta ttattttaaa aaacagcaca aaaggaaact
      841 caccctaact gtaaagtaat tgtgtgtttt gagactataa atatcccttg gagaaaagcc
      901 ttgtttgaat tcctagaccc agctttcttg tacaaagttg gcattaatac gcgttgacat
      961 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     1021 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     1081 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     1141 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     1201 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     1261 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     1321 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     1381 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     1441 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     1501 ggtaggcgtg tacggtggga ggtctatata agcagagctc gtttagtgaa ccgtcagatc
     1561 gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc
     1621 caccggactc tagaggatcg aacccttaag gccaccatgc tgccaccatt ggaaaggctt
     1681 acgctgggga gcgactacaa agacgatgac gacaagggga gcggatccgg ctctatggcg
     1741 tccaatttca ctcagtttgt gctggttgac aacggcggga ccggggacgt tacggtagcc
     1801 ccctcaaact ttgccaacgg tatagcggag tggataagca gcaattctag gagtcaagca
     1861 tacaaagtta catgcagcgt gcgccaatct agcgctcaga atcgcaagta caccattaaa
     1921 gtagaggtcc ccaagggagc ctggagaagc tatcttaaca tggagttgac cataccaatc
     1981 ttcgctacca actctgactg tgaactcatt gtgaaagcca tgcaaggtct gctcaaggat
     2041 ggtaacccaa ttccgtccgc tatcgctgcc aactctggga tttacggggg cagtgggagc
     2101 ggtgcaggat ctggtagtcc agctggggga ggagcaccgg gtagcggtgg ggggagccag
     2161 ctgcacctgc cccaggttct cgcagacgcc gtatcccgcc ttgtaatcgg caagtttggt
     2221 gatcttactg acaatttttc atctcctcat gcgaggcgga tcggcctcgc aggcgtcgtc
     2281 atgacgaccg gaactgacgt gaaagacgcc aaagtcatct gcgtctccac gggcagcaag
     2341 tgcataaacg gggagtacct gagcgaccgg gggctggcac tgaatgattg tcacgctgaa
     2401 atagtgtcta ggcgatctct gcttagattt ctctacactc aactcgaatt gtaccttaac
     2461 aacgaggatg accagaaacg cagtatattt cagaaatcag aacgcggcgg atttcgactt
     2521 aaggaaaaca tccagttcca cttgtatatc agcacatccc cttgcggtga cgcccgaatc
     2581 ttttccccgc acgaggccat attggaggag cccgcctagg cttaagggtt cgatccctac
     2641 tggttagtaa tgagtttaaa cgggggaggc taactgaaac acggaaggag acaataccgg
     2701 aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggg tgttgggtcg
     2761 tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga
     2821 ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg
     2881 ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat agcagatctg
     2941 cgctgatttt gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga
     3001 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
     3061 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca
     3121 ggcttggatc ccaatggcgc gccgagcttg gctcgagcat ggtcatagct gtttcctgtg
     3181 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     3241 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     3301 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     3361 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3421 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3481 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3541 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3601 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3661 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3721 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3781 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3841 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3901 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3961 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     4021 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     4081 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     4141 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     4201 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     4261 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     4321 agttagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca
     4381 ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc
     4441 cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa
     4501 cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg
     4561 actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg ttcaacaggc
     4621 cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat
     4681 tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc
     4741 gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga
     4801 tattcttcta atacctggaa tgctgttttc ccagggatcg cagtggtgag taaccatgca
     4861 tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag
     4921 tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga
     4981 aacaactctg gcgcatcggg cttcccatac aatcgataga ttgtcgcacc tgattgcccg
     5041 acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc
     5101 ggcctagagc aagacgtttc ccgttgaata tggctcatac tcttcctttt tcaatattat
     5161 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     5221 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     5281 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc
     5341 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     5401 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     5461 ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac
     5521 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat
     5581 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
     5641 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
     5701 tcccagtcac gacgttgtaa aacgacggcc agtgaattga cgcgccattg ggat
//
