LOCUS       sequence-337247-        5466 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying a BoxB-MS2 adRNA targeting the RAB7A transcript.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5466)
  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 5466)
  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: BoxB-MS2 adRNA (RAB7A-UAG site)
FEATURES             Location/Qualifiers
     source          1..5466
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     13..35
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     28..44
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     69..85
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   89..229
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   100..229
                     /label=AAV2 ITR
     promoter        248..496
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     248..268
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     419..438
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        539..557
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     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"
     regulatory      1651..1660
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1657..1659,1660..1662,1663..2373)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1657..1659,1660..1662,1663..2373)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1702..1723)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1963..1982)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2310..2331
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2452..2471)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2496..2544
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   2759..2899
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2759..2888
                     /label=AAV2 ITR
     primer_bind     complement(2939..2955)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2939..2955)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2952..2974)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2963..2979
                     /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(2987..2993,2994..3011,3012..3017))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3032..3053
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3341..3929)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3421..3440)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4100..4915)
                     /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(4100..4915)
                     /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     4823..4842
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(4916..5020)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5088..5106
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5144..5166)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5266..5285
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgaattg acgcgccatt
       61 gggatgttgt aaaacgacgg ccagtgaacc tgcaggcagc tgcgcgctcg ctcgctcact
      121 gaggccgccc gggcaaagcc cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc
      181 gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcctg cggccgcacg
      241 cgtggaggag ggcctatttc ccatgattcc ttcatatttg catatacgat acaaggctgt
      301 tagagagata attagaatta atttgactgt aaacacaaag atattagtac aaaatacgtg
      361 acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga
      421 ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat atatcttgtg
      481 gaaaggacga aacaccggcc ctgaaaaagg gccatactgc cgccagctgg atttcccaac
      541 atgaggatca cccatgtctt tttttcgtac 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 ctccggactc tagaggatcg aacccttaag gccaccatgg tgagcaaggg cgaggagctg
     1681 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     1741 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     1801 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     1861 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     1921 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     1981 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     2041 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     2101 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     2161 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     2221 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     2281 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     2341 gggatcactc tcggcatgga cgagctgtac aagtactcag atctcgagct caagtagctt
     2401 aagggttcga tccctactgg ttagtaatga gtttaaacgg gggaggctaa ctgaaacacg
     2461 gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac agaataaaac
     2521 gcacgggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
     2581 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
     2641 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
     2701 ctgccatagc agatctgcgc tgattttgta ggtaaccacg tgcggaccga gcggccgcag
     2761 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     2821 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     2881 gcgcgcagct gcctgcaggc ttggatccca atggcgcgcc gagcttggct cgagcatggt
     2941 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     3001 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     3061 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     3121 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     3181 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     3241 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     3301 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     3361 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     3421 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     3481 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     3541 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     3601 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     3661 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     3721 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     3781 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     3841 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     3901 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     3961 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     4021 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     4081 agtaaacttg gtctgacagt tagaaaaact catcgagcat caaatgaaac tgcaatttat
     4141 tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa
     4201 actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc
     4261 gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga
     4321 aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc atttctttcc
     4381 agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac
     4441 cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg ttaaaaggac
     4501 aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat
     4561 tttcacctga atcaggatat tcttctaata cctggaatgc tgttttccca gggatcgcag
     4621 tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca
     4681 taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac
     4741 ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat cgatagattg
     4801 tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca
     4861 tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg ctcatactct
     4921 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4981 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     5041 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
     5101 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
     5161 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
     5221 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
     5281 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
     5341 accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc
     5401 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
     5461 gggtaa
//
