LOCUS       sequence-472769-        7010 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of ADAR2, sesRNA in mammalian cells with hSyn promoter, 
            with tTA2 as efRNA and ADAR2 overexpression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7010)
  AUTHORS   Qian Y, Li J, Zhao S, Matthews EA, Adoff M, Zhong W, An X, Yeo M, 
            Park C, Yang X, Wang BS, Southwell DG, Huang ZJ
  TITLE     Programmable RNA sensing for cell monitoring and manipulation.
  JOURNAL   Nature. 2022 Oct;610(7933):713-721. doi: 10.1038/s41586-022-05280-1.
            Epub 2022 Oct 5.
  PUBMED    36198803
REFERENCE   2  (bases 1 to 7010)
  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"; doi: 
            "10.1038/s41586-022-05280-1"; journalName: "Nature"; date: 
            "2022-10"; volume: "610"; issue: "7933"; pages: "713-721"
COMMENT     Sequence Label: pAAV-hSyn-ADAR2-2a-Fezf2 sesRNA-2a-tTA2-WPRE
FEATURES             Location/Qualifiers
     source          1..7010
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      77..665
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     566..585
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   727..856
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        901..1348
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      1368..1377
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3477..3533
                     /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             3738..3791
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(3801..4415,4416..4421,4422..4457,4458..4460,4461..4496,
                     4497..4499,4500..4535,4536..4544)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=tTA-Advanced
                     /note="In the Tet-Off(R) system, tTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPAD
                     ALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(4619..4639)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   4999..5139
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4999..5128
                     /label=AAV2 ITR
     rep_origin      5214..5669
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5301..5320)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5511..5532
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5686..5705)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5805..5827
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5865..5883)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5951..6055
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6056..6124,6125..6916)
                     /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     complement(6274..6293)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
       61 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
      121 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      181 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      241 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      301 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      361 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      421 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      481 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      541 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      601 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      661 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      721 acatgtcctg caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg
      781 acctttggtc gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc
      841 atcactaggg gttcctgcgg ccgcacgcgt gtgtctagac tgcagagggc cctgcgtatg
      901 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga
      961 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga
     1021 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg
     1081 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc
     1141 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg
     1201 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg
     1261 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg
     1321 aggagtcgtg tcgtgcctga gagcgcagtc gagaaggtac cggatccgcc accatggata
     1381 tagaagatga agaaaacatg agttccagca gcactgatgt gaaggaaaac cgcaatctgg
     1441 acaacgtgtc ccccaaggat ggcagcacac ctgggcctgg cgagggctct cagctctcca
     1501 atgggggtgg tggtggcccc ggcagaaagc ggcccctgga ggagggcagc aatggccact
     1561 ccaagtaccg cctgaagaaa aggaggaaaa caccagggcc cgtcctcccc aagaacgccc
     1621 tgatgcagct gaatgagatc aagcctggtt tgcagtacac actcctgtcc cagactgggc
     1681 ccgtgcacgc gcctttgttt gtcatgtctg tggaggtgaa tggccaggtt tttgagggct
     1741 ctggtcccac aaagaaaaag gcaaaactcc atgctgctga gaaggccttg aggtctttcg
     1801 ttcagtttcc taatgcctct gaggcccacc tggccatggg gaggaccctg tctgtcaaca
     1861 cggacttcac atctgaccag gccgacttcc ctgacacgct cttcaatggt tttgaaactc
     1921 ctgacaaggc ggagcctccc ttttacgtgg gctccaatgg ggatgactcc ttcagttcca
     1981 gcggggacct cagcttgtct gcttccccgg tgcctgccag cctagcccag cctcctctcc
     2041 ctgtcttacc accattccca cccccgagtg ggaagaatcc cgtgatgatc ttgaacgaac
     2101 tgcgcccagg actcaagtat gacttcctct ccgagagcgg ggagagccat gccaagagct
     2161 tcgtcatgtc tgtggtcgtg gatggtcagt tctttgaagg ctcggggaga aacaagaagc
     2221 ttgccaaggc ccgggctgcg cagtctgccc tggccgccat ttttaacttg cacttggatc
     2281 agacgccatc tcgccagcct attcccagtg agggtcttca gctgcattta ccgcaggttt
     2341 tagctgacgc tgtctcacgc ctggtcctgg gtaagtttgg tgacctgacc gacaacttct
     2401 cctcccctca cgctcgcaga aaagtgctgg ctggagtcgt catgacaaca ggcacagatg
     2461 ttaaagatgc caaggtgata agtgtttcta caggaacaaa atgtattaat ggtgaataca
     2521 tgagtgatcg tggccttgca ttaaatgact gccatgcaga aataatatct cggagatcct
     2581 tgctcagatt tctttataca caacttgagc tttacttaaa taacaaagat gatcaaaaaa
     2641 gatccatctt tcagaaatca gagcgagggg ggtttaggct gaaggagaat gtccagtttc
     2701 atctgtacat cagcacctct ccctgtggag atgccagaat cttctcacca catgagccaa
     2761 tcctggaaga accagcagat agacacccaa atcgtaaagc aagaggacag ctacggacca
     2821 aaatagagtc tggtgagggg acgattccag tgcgctccaa tgcgagcatc caaacgtggg
     2881 acggggtgct gcaaggggag cggctgctca ccatgtcctg cagtgacaag attgcacgct
     2941 ggaacgtggt gggcatccag ggatccctgc tcagcatttt cgtggagccc atttacttct
     3001 cgagcatcat cctgggcagc ctttaccacg gggaccacct ttccagggcc atgtaccagc
     3061 ggatctccaa catagaggac ctgccacctc tctacaccct caacaagcct ttgctcagtg
     3121 gcatcagcaa tgcagaagca cggcagccag ggaaggcccc caacttcagt gtcaactgga
     3181 cggtaggcga ctccgctatt gaggtcatca acgccacgac tgggaaggat gagctgggcc
     3241 gcgcgtcccg cctgtgtaag cacgcgttgt actgtcgctg gatgcgtgtg cacggcaagg
     3301 ttccctccca cttactacgc tccaagatta ccaagcccaa cgtgtaccat gagtccaagc
     3361 tggcggcaaa ggagtaccag gccgccaagg cgcgtctgtt cacagccttc atcaaggcgg
     3421 ggctgggggc ctgggtggag aagcccaccg agcaggacca gttctcactc acgcccgcta
     3481 ctaacttcag cctgctgaag caggctggag acgtggagga gaaccctgga cctgaattcc
     3541 gcagccaggc tggccagttt ggcgttctcc agcagaaaaa gcttggggtg agcagccagg
     3601 gaagtggggg cctgtgcgtt gaggaggccg gataggaaaa ggtggcctcc gaggagctcc
     3661 gaaggtgggt aagcggtgga gtccaggtag ttgaagtagt agagagagcc gctggcaggc
     3721 agccccacgg cgcgccagag ggcagaggaa gtcttctaac atgcggtgac gtggaggaga
     3781 atcccggccc tttaattaac tctagactgg acaagagcaa agtcataaac tctgctctgg
     3841 aattactcaa tgaagtcggt atcgaaggcc tgacgacaag gaaactcgct caaaagctgg
     3901 gagttgagca gcctaccctg tactggcacg tgaagaacaa gcgggccctg ctcgatgccc
     3961 tggcaatcga gatgctggac aggcatcata cccacttctg ccccctggaa ggcgagtcat
     4021 ggcaagactt tctgcggaac aacgccaagt cattccgctg tgctctcctc tcacatcgcg
     4081 acggggctaa agtgcatctc ggcacccgcc caacagagaa acagtacgaa accctggaaa
     4141 atcagctcgc gttcctgtgt cagcaaggct tctccctgga gaacgcactg tacgctctgt
     4201 ccgccgtggg ccactttaca ctgggctgcg tattggagga tcaggagcat caagtagcaa
     4261 aagaggaaag agagacacct accaccgatt ctatgccccc acttctgaga caagcaattg
     4321 agctgttcga ccatcaggga gccgaacctg ccttcctttt cggcctggaa ctaatcatat
     4381 gtggcctgga aaaacagcta aagtgcgaaa gcggcggacc ggccgacgcc cttgacgatt
     4441 ttgacttaga catgctccca gccgatgccc ttgacgactt tgaccttgat atgctgcctg
     4501 ctgacgctct tgacgatttt gaccttgaca tgctccccgg gtaagctagc aattccgctc
     4561 gagataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     4621 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     4681 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttagttctt gccacggcgg
     4741 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     4801 attccgtggt gtttatttgt gaaatttgtg atgctattgc tttatttgta accatctagc
     4861 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     4921 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     4981 gttttttaaa gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
     5041 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     5101 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
     5161 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
     5221 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     5281 tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     5341 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     5401 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     5461 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     5521 gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat
     5581 tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     5641 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
     5701 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     5761 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     5821 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     5881 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     5941 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     6001 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     6061 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     6121 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     6181 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     6241 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     6301 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     6361 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     6421 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     6481 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     6541 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     6601 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     6661 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     6721 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     6781 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     6841 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     6901 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     6961 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
//
