LOCUS       sequence-337248-        5477 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying a MS2 adRNA targeting the RAB7A transcript.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5477)
  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 5477)
  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: MS2-adRNA (RAB7A-GAC site)
FEATURES             Location/Qualifiers
     source          1..5477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   110..250
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   110..239
                     /label=AAV2 ITR
     primer_bind     complement(290..306)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(290..306)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(303..325)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    314..330
                     /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(338..344,345..362,363..368))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    383..404
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(692..1280)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(772..791)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1451..2266)
                     /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(1451..2266)
                     /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     2174..2193
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2267..2371)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2439..2457
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2495..2517)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2617..2636
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2830..2852
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2845..2861
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2886..2902
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   2906..3046
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2917..3046
                     /label=AAV2 ITR
     promoter        3065..3313
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     3065..3085
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3236..3255
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        3316..3334
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        3367..3385
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        3420..3733
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     3652..3672
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(3745..3769)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        3784..4163
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4164..4367
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4318..4338
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4364..4388
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      4479..4488
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4485..4487,4488..4490,4491..5201)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4485..4487,4488..4490,4491..5201)
                     /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(4530..4551)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4791..4810)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5138..5159
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(5280..5299)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5324..5372
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag
       61 cagatctgcg ctgattttgt aggtaaccac gtgcggaccg agcggccgca ggaaccccta
      121 gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca
      181 aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc
      241 tgcctgcagg cttggatccc aatggcgcgc cgagcttggc tcgagcatgg tcatagctgt
      301 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
      361 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
      421 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
      481 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
      541 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
      601 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
      661 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
      721 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
      781 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
      841 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
      901 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
      961 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     1021 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     1081 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     1141 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     1201 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     1261 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     1321 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     1381 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     1441 ggtctgacag ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag
     1501 gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga
     1561 ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat
     1621 caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat
     1681 gagtgacgac tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt
     1741 caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca
     1801 ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa
     1861 caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg
     1921 aatcaggata ttcttctaat acctggaatg ctgttttccc agggatcgca gtggtgagta
     1981 accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg
     2041 tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat
     2101 gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg
     2161 attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat
     2221 ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg gctcatactc ttcctttttc
     2281 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     2341 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     2401 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     2461 ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     2521 cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag
     2581 cgggtgttgg cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga
     2641 gagtgcacca tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca
     2701 ggcgccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     2761 cgctattacg ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc
     2821 cagggttttc ccagtcacga cgttgtaaaa cgacggccag tgaattgacg cgccattggg
     2881 atgttgtaaa acgacggcca gtgaacctgc aggcagctgc gcgctcgctc gctcactgag
     2941 gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag
     3001 cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctgcgg ccgcacgcgt
     3061 ggaggagggc ctatttccca tgattccttc atatttgcat atacgataca aggctgttag
     3121 agagataatt agaattaatt tgactgtaaa cacaaagata ttagtacaaa atacgtgacg
     3181 tagaaagtaa taatttcttg ggtagtttgc agttttaaaa ttatgtttta aaatggacta
     3241 tcatatgctt accgtaactt gaaagtattt cgatttcttg gctttatata tcttgtggaa
     3301 aggacgaaac accgaacatg aggatcaccc atgtctacat aattcttgtg cctactgtac
     3361 agaataacat gaggatcacc catgtctttt tttcgtactg agtcgcccag tctcagatag
     3421 atccgacgcc gccatctcta ggcccgcgcc ggccccctcg cacagacttg tgggagaagc
     3481 tcggctactc ccctgccccg gttaatttgc atataatatt tcctagtaac tatagaggct
     3541 taatgtgcga taaaagacag ataatctgtt ctttttaata ctagctacat tttacatgat
     3601 aggcttggat ttctataaga gatacaaata ctaaattatt attttaaaaa acagcacaaa
     3661 aggaaactca ccctaactgt aaagtaattg tgtgttttga gactataaat atcccttgga
     3721 gaaaagcctt gtttgaattc ctagacccag ctttcttgta caaagttggc attaatacgc
     3781 gttgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
     3841 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
     3901 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
     3961 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     4021 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     4081 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     4141 tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat
     4201 agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt
     4261 tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc
     4321 aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt ttagtgaacc
     4381 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
     4441 gatccagcct ccggactcta gaggatcgaa cccttaaggc caccatggtg agcaagggcg
     4501 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     4561 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     4621 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga
     4681 cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca
     4741 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     4801 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     4861 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     4921 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     4981 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     5041 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt
     5101 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     5161 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtactcagat ctcgagctca
     5221 agtagcttaa gggttcgatc cctactggtt agtaatgagt ttaaacgggg gaggctaact
     5281 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     5341 aataaaacgc acgggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg
     5401 gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt
     5461 ttccccaccc caccccc
//
