LOCUS       sequence-336848-        7047 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying ADAR2-DDC(E488Q)-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)
FEATURES             Location/Qualifiers
     source          1..7047
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        136..384
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     136..156
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     307..326
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        386..460
                     /label=Cj gRNA scaffold
                     /note="guide RNA scaffold for the Campylobacter jejuni 
                     CRISPR/Cas9 system (Kim et al., 2016)"
     promoter        545..858
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     777..797
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(870..894)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        909..1288
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1289..1492
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1443..1463
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1489..1513
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1604..1613
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2777..2803
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2810..2836
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             2846..2902
                     /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             2906..2932
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             2939..2962
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3921..3940)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3965..4013
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   4228..4368
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4228..4357
                     /label=AAV2 ITR
     primer_bind     complement(4408..4424)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4408..4424)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4421..4443)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4432..4448
                     /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(4456..4462,4463..4480,4481..4486))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4501..4522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4810..5398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4890..4909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5569..6384)
                     /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(5569..6384)
                     /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     6292..6311
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(6385..6489)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6557..6575
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6613..6635)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6735..6754
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6948..6970
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6963..6979
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7004..7020
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   join(7024..7047,1..117)
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   join(7035..7047,1..117)
                     /label=AAV2 ITR
ORIGIN
        1 cgctcactga ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt cgcccggcct
       61 cagtgagcga gcgagcgcgc agagagggag tggccaactc catcactagg ggttcctgcg
      121 gccgcacgcg tggaggaggg cctatttccc atgattcctt catatttgca tatacgatac
      181 aaggctgtta gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa
      241 aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt
      301 aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat
      361 atcttgtgga aaggacgaaa caccggtttt agtccctgaa ggaagggact aaaataaaga
      421 gtttgcggga ctctgcgggg ttacaatccc ctaaaaccgc taaggctagt ccgttatcaa
      481 cttgaaaaag tggcaccgag tcggtgcttt tttgctagcg tactgagtcg cccagtctca
      541 gatagatccg acgccgccat ctctaggccc gcgccggccc cctcgcacag acttgtggga
      601 gaagctcggc tactcccctg ccccggttaa tttgcatata atatttccta gtaactatag
      661 aggcttaatg tgcgataaaa gacagataat ctgttctttt taatactagc tacattttac
      721 atgataggct tggatttcta taagagatac aaatactaaa ttattatttt aaaaaacagc
      781 acaaaaggaa actcacccta actgtaaagt aattgtgtgt tttgagacta taaatatccc
      841 ttggagaaaa gccttgtttg aattcctaga cccagctttc ttgtacaaag ttggcattaa
      901 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      961 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     1021 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     1081 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     1141 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     1201 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     1261 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     1321 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     1381 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     1441 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctcgtttagt
     1501 gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata gaagacaccg
     1561 ggaccgatcc agcctccgga ctctagagga tcgaaccctt aaggccacca tggacagaca
     1621 tcctaatagg aaggctagag gccaacttcg gacgaagatt gaaagtggcc agggtactat
     1681 cccggtgcgg tccaacgcta gtattcaaac gtgggacgga gtccttcaag gtgaacggct
     1741 gttgacaatg agctgctcag acaaaatcgc gcgctggaat gtagtgggaa tccaaggcag
     1801 cctcttgagc atattcgtag aacccatata tttctcatcc attattttgg gctctctgta
     1861 tcatggtgac catctgtcaa gggctatgta ccaacgaatt tctaatatcg aggatcttcc
     1921 tccactctat acactcaata agcctctctt gtccgggata tcaaacgctg aggcccgcca
     1981 gccagggaaa gctcctaact tcagtgttaa ctggaccgtt ggtgattctg cgatagaggt
     2041 catcaacgcc acgacaggta aggatgagct cggtagagcc tcacgcctgt gtaaacacgc
     2101 gttgtattgt agatggatga gagtacatgg gaaggtccca tctcacttgc tccgaagcaa
     2161 gatcactaag cctaatgtgt atcatgagtc aaaactcgcg gctaaagaat accaggcagc
     2221 caaagctcga ctttttacag cttttattaa ggcagggctc ggggcatggg tcgagaagcc
     2281 gaccgagcag gaccaattct ctctgacgcc cgggggcagt gggagcggtg caggatctgg
     2341 tagtccagct gggggaggag caccgggtag cggtgggggg agcaatgcga ggacacggcg
     2401 acgagaacga cgcgctgaaa agcaagcgca atggaaagct gcaaacggag gtggaggatc
     2461 tggcggtgga gggtcaggtg gcggcggttc aaatgctcgg actcggagac gcgagcggag
     2521 agcggagaag caggcacaat ggaaagccgc taatggtggc ggcgggagtg ggggaggcgg
     2581 atctggaggt ggcgggtcca acgcgagaac ccgccgacga gagcgcagag cggagaaaca
     2641 agcacagtgg aaggcggcca acgggggtgg gggatctggc ggcgggggat ccggcggagg
     2701 agggtccaat gcccgaaccc gacgaaggga aagacgggct gagaaacagg ctcaatggaa
     2761 agcggctaat gggagctacc catacgatgt tccagattac gctggatccc tgccaccatt
     2821 ggaaagactg accctcggat ccggcgcaac aaacttctct ctgctgaaac aagccggaga
     2881 tgtcgaagag aatcctggac cgatgctgcc tcccctcgaa aggcttacgc tggggagcga
     2941 ctacaaagac gatgacgaca aggggagcgg atccggctct atggcgtcca atttcactca
     3001 gtttgtgctg gttgacaacg gcgggaccgg ggacgttacg gtagccccct caaactttgc
     3061 caacggtata gcggagtgga taagcagcaa ttctaggagt caagcataca aagttacatg
     3121 cagcgtgcgc caatctagcg ctcagaatcg caagtacacc attaaagtag aggtccccaa
     3181 gggagcctgg agaagctatc ttaacatgga gttgaccata ccaatcttcg ctaccaactc
     3241 tgactgtgaa ctcattgtga aagccatgca aggtctgctc aaggatggta acccaattcc
     3301 gtccgctatc gctgccaact ctgggattta cgggggcagt gggagcggtg caggatctgg
     3361 tagtccagct gggggaggag caccgggtag cggtgggggg agccagctgc acctgcccca
     3421 ggttctcgca gacgccgtat cccgccttgt actgggcaag tttggtgatc ttactgacaa
     3481 tttttcatct cctcatgcga ggcggaaagt actcgcaggc gtcgtcatga cgaccggaac
     3541 tgacgtgaaa gacgccaaag tcatctctgt ctccacgggc acaaagtgca taaacgggga
     3601 gtacatgagc gaccgggggc tggcactgaa tgattgtcac gctgaaataa tatctaggcg
     3661 atctctgctt agatttctct acactcaact cgaattgtac cttaacaaca aagatgacca
     3721 gaaacgcagt atatttcaga aatcagaacg cggcggattt cgacttaagg aaaacgttca
     3781 gttccacttg tatatcagca catccccttg cggtgacgcc cgaatctttt ccccgcacga
     3841 gccgatattg gaggagcccg cgtgagctta agggttcgat ccctactggt tagtaatgag
     3901 tttaaacggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     3961 atgacggcaa taaaaagaca gaataaaacg cacgggtgtt gggtcgtttg ttcataaacg
     4021 cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca
     4081 atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg
     4141 gctcgcagcc aacgtcgggg cggcaggccc tgccatagca gatctgcgct gattttgtag
     4201 gtaaccacgt gcggaccgag cggccgcagg aacccctagt gatggagttg gccactccct
     4261 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     4321 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcaggct tggatcccaa
     4381 tggcgcgccg agcttggctc gagcatggtc atagctgttt cctgtgtgaa attgttatcc
     4441 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4501 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4561 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     4621 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4681 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4741 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4801 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4861 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4921 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4981 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5041 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5101 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5161 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5221 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     5281 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5341 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5401 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5461 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5521 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt agaaaaactc
     5581 atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg
     5641 aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag
     5701 atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc
     5761 ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga
     5821 gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc cattacgctc
     5881 gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag
     5941 acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg
     6001 caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac
     6061 ctggaatgct gttttcccag ggatcgcagt ggtgagtaac catgcatcat caggagtacg
     6121 gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat
     6181 ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc
     6241 atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc
     6301 ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc tagagcaaga
     6361 cgtttcccgt tgaatatggc tcatactctt cctttttcaa tattattgaa gcatttatca
     6421 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6481 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6541 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga
     6601 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     6661 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     6721 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga
     6781 aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc cattcaggct
     6841 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     6901 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     6961 ttgtaaaacg acggccagtg aattgacgcg ccattgggat gttgtaaaac gacggccagt
     7021 gaacctgcag gcagctgcgc gctcgct
//
