LOCUS       sequence-337250-        7047 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying ADAR2-DDC(E488Q, 
            N496F)-lambda-N-P2A-MCP-ADAR2-DDN with a nuclear export signal.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7047)
  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 7047)
  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: All-in-one split-ADAR2(E488Q, N496F)
FEATURES             Location/Qualifiers
     source          1..7047
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        138..386
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     138..158
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     309..328
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        388..462
                     /label=Cj gRNA scaffold
                     /note="guide RNA scaffold for the Campylobacter jejuni 
                     CRISPR/Cas9 system (Kim et al., 2016)"
     promoter        547..860
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     779..799
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(872..896)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        911..1290
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1291..1494
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1445..1465
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1491..1515
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1606..1615
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2779..2805
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2812..2838
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             2848..2904
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2908..2934
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             2941..2964
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3923..3942)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3967..4015
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   4230..4370
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4230..4359
                     /label=AAV2 ITR
     primer_bind     complement(4410..4426)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4410..4426)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4423..4445)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4434..4450
                     /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(4458..4464,4465..4482,4483..4488))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4503..4524
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4812..5400)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4892..4911)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5571..6386)
                     /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(5571..6386)
                     /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     6294..6313
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(6387..6491)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6559..6577
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6615..6637)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6737..6756
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6950..6972
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6965..6981
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7006..7022
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   join(7026..7047,1..119)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   join(7037..7047,1..119)
                     /label=AAV2 ITR
ORIGIN
        1 ctcgctcact gaggccgccc gggcaaagcc cgggcgtcgg gcgacctttg gtcgcccggc
       61 ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcctg
      121 cggccgcacg cgtggaggag ggcctatttc ccatgattcc ttcatatttg catatacgat
      181 acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag atattagtac
      241 aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt
      301 ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat
      361 atatcttgtg gaaaggacga aacaccggtt ttagtccctg aaggaaggga ctaaaataaa
      421 gagtttgcgg gactctgcgg ggttacaatc ccctaaaacc gctaaggcta gtccgttatc
      481 aacttgaaaa agtggcaccg agtcggtgct tttttgctag cgtactgagt cgcccagtct
      541 cagatagatc cgacgccgcc atctctaggc ccgcgccggc cccctcgcac agacttgtgg
      601 gagaagctcg gctactcccc tgccccggtt aatttgcata taatatttcc tagtaactat
      661 agaggcttaa tgtgcgataa aagacagata atctgttctt tttaatacta gctacatttt
      721 acatgatagg cttggatttc tataagagat acaaatacta aattattatt ttaaaaaaca
      781 gcacaaaagg aaactcaccc taactgtaaa gtaattgtgt gttttgagac tataaatatc
      841 ccttggagaa aagccttgtt tgaattccta gacccagctt tcttgtacaa agttggcatt
      901 aatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
      961 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
     1021 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
     1081 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     1141 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     1201 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1261 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     1321 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     1381 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     1441 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta
     1501 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac
     1561 cgggaccgat ccagcctccg gactctagag gatcgaaccc ttaaggccac catggacaga
     1621 catcctaata ggaaggctag aggccaactt cggacgaaga ttgaaagtgg ccagggtact
     1681 atcccggtgc ggtccttcgc tagtattcaa acgtgggacg gagtccttca aggtgaacgg
     1741 ctgttgacaa tgagctgctc agacaaaatc gcgcgctgga atgtagtggg aatccaaggc
     1801 agcctcttga gcatattcgt agaacccata tatttctcat ccattatttt gggctctctg
     1861 tatcatggtg accatctgtc aagggctatg taccaacgaa tttctaatat cgaggatctt
     1921 cctccactct atacactcaa taagcctctc ttgtccggga tatcaaacgc tgaggcccgc
     1981 cagccaggga aagctcctaa cttcagtgtt aactggaccg ttggtgattc tgcgatagag
     2041 gtcatcaacg ccacgacagg taaggatgag ctcggtagag cctcacgcct gtgtaaacac
     2101 gcgttgtatt gtagatggat gagagtacat gggaaggtcc catctcactt gctccgaagc
     2161 aagatcacta agcctaatgt gtatcatgag tcaaaactcg cggctaaaga ataccaggca
     2221 gccaaagctc gactttttac agcttttatt aaggcagggc tcggggcatg ggtcgagaag
     2281 ccgaccgagc aggaccaatt ctctctgacg cccgggggca gtgggagcgg tgcaggatct
     2341 ggtagtccag ctgggggagg agcaccgggt agcggtgggg ggagcaatgc gaggacacgg
     2401 cgacgagaac gacgcgctga aaagcaagcg caatggaaag ctgcaaacgg aggtggagga
     2461 tctggcggtg gagggtcagg tggcggcggt tcaaatgctc ggactcggag acgcgagcgg
     2521 agagcggaga agcaggcaca atggaaagcc gctaatggtg gcggcgggag tgggggaggc
     2581 ggatctggag gtggcgggtc caacgcgaga acccgccgac gagagcgcag agcggagaaa
     2641 caagcacagt ggaaggcggc caacgggggt gggggatctg gcggcggggg atccggcgga
     2701 ggagggtcca atgcccgaac ccgacgaagg gaaagacggg ctgagaaaca ggctcaatgg
     2761 aaagcggcta atgggagcta cccatacgat gttccagatt acgctggatc cctgccacca
     2821 ttggaaagac tgaccctcgg atccggcgca acaaacttct ctctgctgaa acaagccgga
     2881 gatgtcgaag agaatcctgg accgatgctg cctcccctcg aaaggcttac gctggggagc
     2941 gactacaaag acgatgacga caaggggagc ggatccggct ctatggcgtc caatttcact
     3001 cagtttgtgc tggttgacaa cggcgggacc ggggacgtta cggtagcccc ctcaaacttt
     3061 gccaacggta tagcggagtg gataagcagc aattctagga gtcaagcata caaagttaca
     3121 tgcagcgtgc gccaatctag cgctcagaat cgcaagtaca ccattaaagt agaggtcccc
     3181 aagggagcct ggagaagcta tcttaacatg gagttgacca taccaatctt cgctaccaac
     3241 tctgactgtg aactcattgt gaaagccatg caaggtctgc tcaaggatgg taacccaatt
     3301 ccgtccgcta tcgctgccaa ctctgggatt tacgggggca gtgggagcgg tgcaggatct
     3361 ggtagtccag ctgggggagg agcaccgggt agcggtgggg ggagccagct gcacctgccc
     3421 caggttctcg cagacgccgt atcccgcctt gtactgggca agtttggtga tcttactgac
     3481 aatttttcat ctcctcatgc gaggcggaaa gtactcgcag gcgtcgtcat gacgaccgga
     3541 actgacgtga aagacgccaa agtcatctct gtctccacgg gcacaaagtg cataaacggg
     3601 gagtacatga gcgaccgggg gctggcactg aatgattgtc acgctgaaat aatatctagg
     3661 cgatctctgc ttagatttct ctacactcaa ctcgaattgt accttaacaa caaagatgac
     3721 cagaaacgca gtatatttca gaaatcagaa cgcggcggat ttcgacttaa ggaaaacgtt
     3781 cagttccact tgtatatcag cacatcccct tgcggtgacg cccgaatctt ttccccgcac
     3841 gagccgatat tggaggagcc cgcgtgagct taagggttcg atccctactg gttagtaatg
     3901 agtttaaacg ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg
     3961 ctatgacggc aataaaaaga cagaataaaa cgcacgggtg ttgggtcgtt tgttcataaa
     4021 cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc
     4081 caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca
     4141 gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cagatctgcg ctgattttgt
     4201 aggtaaccac gtgcggaccg agcggccgca ggaaccccta gtgatggagt tggccactcc
     4261 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     4321 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg cttggatccc
     4381 aatggcgcgc cgagcttggc tcgagcatgg tcatagctgt ttcctgtgtg aaattgttat
     4441 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     4501 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     4561 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     4621 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4681 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4741 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4801 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4861 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4921 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4981 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     5041 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     5101 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     5161 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     5221 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     5281 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     5341 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     5401 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     5461 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     5521 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttagaaaaac
     5581 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
     5641 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
     5701 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
     5761 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
     5821 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
     5881 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
     5941 agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
     6001 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
     6061 acctggaatg ctgttttccc agggatcgca gtggtgagta accatgcatc atcaggagta
     6121 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
     6181 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
     6241 gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga
     6301 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa
     6361 gacgtttccc gttgaatatg gctcatactc ttcctttttc aatattattg aagcatttat
     6421 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6481 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc
     6541 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt
     6601 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
     6661 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
     6721 ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt
     6781 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc gccattcagg
     6841 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg
     6901 aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga
     6961 cgttgtaaaa cgacggccag tgaattgacg cgccattggg atgttgtaaa acgacggcca
     7021 gtgaacctgc aggcagctgc gcgctcg
//
