LOCUS       sequence-337168-        5781 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying a guide RNA targeting the human RAB7A mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5781)
  AUTHORS   Katrekar D, Yen J, Xiang Y, Saha A, Meluzzi D, Savva Y, Mali P
  TITLE     Efficient in vitro and in vivo RNA editing via recruitment of 
            endogenous ADARs using circular guide RNAs.
  JOURNAL   Nat Biotechnol. 2022 Feb 10. pii: 10.1038/s41587-021-01171-4. doi: 
            10.1038/s41587-021-01171-4.
  PUBMED    35145312
REFERENCE   2  (bases 1 to 5781)
  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: "Nat 
            Biotechnol. 2022 Feb 10. pii: 10.1038/s41587-021-01171-4. doi: 
            10.1038/s41587-021-01171-4."
COMMENT     Sequence Label: Circular 200,100 (RAB7A)
FEATURES             Location/Qualifiers
     source          1..5781
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(283..871)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(363..382)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1042..1857)
                     /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(1042..1857)
                     /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     1765..1784
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1858..1962)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2030..2048
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2086..2108)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2208..2227
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2421..2443
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2436..2452
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2477..2493
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   2497..2637
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2508..2637
                     /label=AAV2 ITR
     promoter        2656..2904
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2656..2676
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2827..2846
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        3315..3628
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     3547..3567
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(3640..3664)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        3679..4058
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4059..4262
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4213..4233
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4259..4283
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      4374..4383
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4380..4382,4383..4385,4386..5096)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4380..4382,4383..4385,4386..5096)
                     /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(4425..4446)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4686..4705)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5033..5054
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(5175..5194)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5219..5267
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   5482..5622
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5482..5611
                     /label=AAV2 ITR
     primer_bind     complement(5662..5678)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5662..5678)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5675..5697)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5686..5702
                     /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(5710..5716,5717..5734,5735..5740))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5755..5776
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
       61 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
      121 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
      181 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
      241 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
      301 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
      361 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
      421 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
      481 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
      541 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
      601 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
      661 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
      721 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
      781 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
      841 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
      901 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
      961 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     1021 tgagtaaact tggtctgaca gttagaaaaa ctcatcgagc atcaaatgaa actgcaattt
     1081 attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta atgaaggaga
     1141 aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg cgattccgac
     1201 tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt tatcaagtga
     1261 gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat gcatttcttt
     1321 ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg catcaaccaa
     1381 accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg
     1441 acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg catcaacaat
     1501 attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc cagggatcgc
     1561 agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg tcggaagagg
     1621 cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat tggcaacgct
     1681 acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca atcgatagat
     1741 tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata aatcagcatc
     1801 catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat ggctcatact
     1861 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     1921 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     1981 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     2041 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     2101 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     2161 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
     2221 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     2281 ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     2341 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     2401 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattgac
     2461 gcgccattgg gatgttgtaa aacgacggcc agtgaacctg caggcagctg cgcgctcgct
     2521 cgctcactga ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt cgcccggcct
     2581 cagtgagcga gcgagcgcgc agagagggag tggccaactc catcactagg ggttcctgcg
     2641 gccgcacgcg tggaggaggg cctatttccc atgattcctt catatttgca tatacgatac
     2701 aaggctgtta gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa
     2761 aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt
     2821 aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat
     2881 accttgtgga aaggacgaaa caccgccatc agtcgccggt cccaagcccg gataaaatgg
     2941 gagggggcgg gaaaccgcct aaccatgccg actgatggca gaaaaaaaaa agacagttgt
     3001 ccccctggag agatgaaaag cttgttggct cttaagtctt tgataaaagg cgtacataat
     3061 tcttgtgtct actgtacaga atactgccgc cagctggatt tcccaattct gagtaacact
     3121 ctgcaatcca aacagggttc aaccctccac cttacaggcc tgcattacag gacttaaaca
     3181 cataatccaa aaaaaaaaaa ctgccatcag tcggcgtgga ctgtagaaca ctgccaatgc
     3241 cggtcccaag cccggataaa agtggagggt acagtccacg cttttttgcg tactgagtcg
     3301 cccagtctca gatagatccg acgccgccat ctctaggccc gcgccggccc cctcgcacag
     3361 acttgtggga gaagctcggc tactcccctg ccccggttaa tttgcatata atatttccta
     3421 gtaactatag aggcttaatg tgcgataaaa gacagataat ctgttctttt taatactagc
     3481 tacattttac atgataggct tggatttcta taagagatac aaatactaaa ttattatttt
     3541 aaaaaacagc acaaaaggaa actcacccta actgtaaagt aattgtgtgt tttgagacta
     3601 taaatatccc ttggagaaaa gccttgtttg aattcctaga cccagctttc ttgtacaaag
     3661 ttggcattaa tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg
     3721 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     3781 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     3841 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     3901 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     3961 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     4021 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     4081 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     4141 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     4201 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     4261 ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata
     4321 gaagacaccg ggaccgatcc agcctccgga ctctagagga tcgaaccctt aaggccacca
     4381 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     4441 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     4501 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     4561 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     4621 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     4681 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     4741 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     4801 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     4861 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     4921 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     4981 acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     5041 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtact
     5101 cagatctcga gctcaagtag cttaagggtt cgatccctac tggttagtaa tgagtttaaa
     5161 cgggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     5221 gcaataaaaa gacagaataa aacgcacggg tgttgggtcg tttgttcata aacgcggggt
     5281 tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc
     5341 ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc
     5401 agccaacgtc ggggcggcag gccctgccat agcagatctg cgctgatttt gtaggtaacc
     5461 acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     5521 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     5581 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggcttggatc ccaatggcgc
     5641 gccgagcttg gctcgagcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5701 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5761 gagctaactc acattaattg c
//
