LOCUS       sequence-244437-        6900 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses adRNA(1,20,6) targeting the RAB7A transcript along with 
            the ADAR2(E488Q).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6900)
  AUTHORS   Katrekar D, Chen G, Meluzzi D, Ganesh A, Worlikar A, Shih YR, 
            Varghese S, Mali P
  TITLE     In vivo RNA editing of point mutations via RNA-guided adenosine 
            deaminases.
  JOURNAL   Nat Methods. 2019 Mar;16(3):239-242. doi: 10.1038/s41592-019-0323-0.
            Epub 2019 Feb 8.
  PUBMED    30737497
REFERENCE   2  (bases 1 to 6900)
  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/s41592-019-0323-0"; journalName: "Nat Methods"; date: 
            "2019-03"; volume: "16"; issue: "3"; pages: "239-242"
COMMENT     Sequence Label: GluR2_adRNA(1,20,6)-ADAR2(E488Q)
FEATURES             Location/Qualifiers
     source          1..6900
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(1..589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(81..100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(760..1575)
                     /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(760..1575)
                     /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     1483..1502
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1576..1680)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1748..1766
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1804..1826)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1926..1945
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2139..2161
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2154..2170
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2195..2211
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   2215..2355
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2226..2355
                     /label=AAV2 ITR
     promoter        2374..2622
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2374..2394
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2545..2564
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        2720..3033
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2952..2972
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(3112..3136)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        3151..3530
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3531..3734
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3685..3705
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3731..3755
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      3846..3855
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(6012..6031)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6056..6104
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   6319..6459
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6319..6448
                     /label=AAV2 ITR
     primer_bind     complement(6499..6515)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6499..6515)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6512..6534)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6523..6539
                     /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(6547..6553,6554..6571,6572..6577))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6592..6613
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
       61 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
      121 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
      181 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
      241 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
      301 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
      361 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
      421 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
      481 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
      541 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
      601 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
      661 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
      721 atcaatctaa agtatatatg agtaaacttg gtctgacagt tagaaaaact catcgagcat
      781 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
      841 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
      901 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
      961 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     1021 aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
     1081 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac
     1141 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
     1201 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
     1261 tgttttccca gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
     1321 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
     1381 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
     1441 cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
     1501 cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg
     1561 ttgaatatgg ctcatactct tcctttttca atattattga agcatttatc agggttattg
     1621 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     1681 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     1741 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     1801 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     1861 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
     1921 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
     1981 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
     2041 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     2101 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     2161 gacggccagt gaattgacgc gccattggga tgttgtaaaa cgacggccag tgaacctgca
     2221 ggcagctgcg cgctcgctcg ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga
     2281 cctttggtcg cccggcctca gtgagcgagc gagcgcgcag agagggagtg gccaactcca
     2341 tcactagggg ttcctgcggc cgcacgcgtg gaggagggcc tatttcccat gattccttca
     2401 tatttgcata tacgatacaa ggctgttaga gagataatta gaattaattt gactgtaaac
     2461 acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca
     2521 gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc
     2581 gatttcttgg ctttatatat cttgtggaaa ggacgaaaca ccgtggaata gtataacaat
     2641 atgctaaatg ttgttatagt atcccactgc cgccagctgg atttcccttt tttcgtactg
     2701 agtcgcccag tctcagatag atccgacgcc gccatctcta ggcccgcgcc ggccccctcg
     2761 cacagacttg tgggagaagc tcggctactc ccctgccccg gttaatttgc atataatatt
     2821 tcctagtaac tatagaggct taatgtgcga taaaagacag ataatctgtt ctttttaata
     2881 ctagctacat tttacatgat aggcttggat ttctataaga gatacaaata ctaaattatt
     2941 attttaaaaa acagcacaaa aggaaactca ccctaactgt aaagtaattg tgtgttttga
     3001 gactataaat atcccttgga gaaaagcctt gtttggtgga atagtataac aatatgctaa
     3061 atgttgttat agtatcccac tgccgccagc tggatttccc ttttttccta gacccagctt
     3121 tcttgtacaa agttggcatt aatacgcgtt gacattgatt attgactagt tattaatagt
     3181 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     3241 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     3301 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     3361 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     3421 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     3481 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     3541 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     3601 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     3661 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     3721 atataagcag agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
     3781 ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagag gatcgaaccc
     3841 ttaaggccac catggatata gaagatgaag aaaacatgag ttccagcagc actgatgtga
     3901 aggaaaaccg caatctggac aacgtgtccc ccaaggatgg cagcacacct gggcctggcg
     3961 agggctctca gctctccaat gggggtggtg gtggccccgg cagaaagcgg cccctggagg
     4021 agggcagcaa tggccactcc aagtaccgcc tgaagaaaag gaggaaaaca ccagggcccg
     4081 tcctccccaa gaacgccctg atgcagctga atgagatcaa gcctggtttg cagtacacac
     4141 tcctgtccca gactgggccc gtgcacgcgc ctttgtttgt catgtctgtg gaggtgaatg
     4201 gccaggtttt tgagggctct ggtcccacaa agaaaaaggc aaaactccat gctgctgaga
     4261 aggccttgag gtctttcgtt cagtttccta atgcctctga ggcccacctg gccatgggga
     4321 ggaccctgtc tgtcaacacg gacttcacat ctgaccaggc cgacttccct gacacgctct
     4381 tcaatggttt tgaaactcct gacaaggcgg agcctccctt ttacgtgggc tccaatgggg
     4441 atgactcctt cagttccagc ggggacctca gcttgtctgc ttccccggtg cctgccagcc
     4501 tagcccagcc tcctctccct gtcttaccac cattcccacc cccgagtggg aagaatcccg
     4561 tgatgatctt gaacgaactg cgcccaggac tcaagtatga cttcctctcc gagagcgggg
     4621 agagccatgc caagagcttc gtcatgtctg tggtcgtgga tggtcagttc tttgaaggct
     4681 cggggagaaa caagaagctt gccaaggccc gggctgcgca gtctgccctg gccgccattt
     4741 ttaacttgca cttggatcag acgccatctc gccagcctat tcccagtgag ggtcttcagc
     4801 tgcatttacc gcaggtttta gctgacgctg tctcacgcct ggtcctgggt aagtttggtg
     4861 acctgaccga caacttctcc tcccctcacg ctcgcagaaa agtgctggct ggagtcgtca
     4921 tgacaacagg cacagatgtt aaagatgcca aggtgataag tgtttctaca ggaacaaaat
     4981 gtattaatgg tgaatacatg agtgatcgtg gccttgcatt aaatgactgc catgcagaaa
     5041 taatatctcg gagatccttg ctcagatttc tttatacaca acttgagctt tacttaaata
     5101 acaaagatga tcaaaaaaga tccatctttc agaaatcaga gcgagggggg tttaggctga
     5161 aggagaatgt ccagtttcat ctgtacatca gcacctctcc ctgtggagat gccagaatct
     5221 tctcaccaca tgagccaatc ctggaagaac cagcagatag acacccaaat cgtaaagcaa
     5281 gaggacagct acggaccaaa atagagtctg gtcaggggac gattccagtg cgctccaatg
     5341 cgagcatcca aacgtgggac ggggtgctgc aaggggagcg gctgctcacc atgtcctgca
     5401 gtgacaagat tgcacgctgg aacgtggtgg gcatccaggg atccctgctc agcattttcg
     5461 tggagcccat ttacttctcg agcatcatcc tgggcagcct ttaccacggg gaccaccttt
     5521 ccagggccat gtaccagcgg atctccaaca tagaggacct gccacctctc tacaccctca
     5581 acaagccttt gctcagtggc atcagcaatg cagaagcacg gcagccaggg aaggccccca
     5641 acttcagtgt caactggacg gtaggcgact ccgctattga ggtcatcaac gccacgactg
     5701 ggaaggatga gctgggccgc gcgtcccgcc tgtgtaagca cgcgttgtac tgtcgctgga
     5761 tgcgtgtgca cggcaaggtt ccctcccact tactacgctc caagattacc aagcccaacg
     5821 tgtaccatga gtccaagctg gcggcaaagg agtaccaggc cgccaaggcg cgtctgttca
     5881 cagccttcat caaggcgggg ctgggggcct gggtggagaa gcccaccgag caggaccagt
     5941 tctcactcac gccctgactt aagggttcga tccctactgg ttagtaatga gtttaaacgg
     6001 gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca
     6061 ataaaaagac agaataaaac gcacgggtgt tgggtcgttt gttcataaac gcggggttcg
     6121 gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc aatacgcccg
     6181 cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag ggctcgcagc
     6241 caacgtcggg gcggcaggcc ctgccatagc agatctgcgc tgattttgta ggtaaccacg
     6301 tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
     6361 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     6421 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggc ttggatccca atggcgcgcc
     6481 gagcttggct cgagcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     6541 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     6601 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     6661 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     6721 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     6781 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     6841 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
//
