LOCUS       sequence-418382-        3635 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector expressing tracrRNA under U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3635)
  AUTHORS   Zhang H, Kelly K, Lee J, Echeverria D, Cooper D, Panwala R, Amrani 
            N, Chen Z, Gaston N, Wagh A, Newby GA, Xie J, Liu DR, Gao G, Wolfe 
            SA, Khvorova A, Watts JK, Sontheimer EJ
  TITLE     Self-delivering, chemically modified CRISPR RNAs for AAV co-delivery
            and genome editing in vivo.
  JOURNAL   Nucleic Acids Res. 2023 Nov 30:gkad1125. doi: 10.1093/nar/gkad1125.
  PUBMED    38033325
REFERENCE   2  (bases 1 to 3635)
  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: "Nucleic 
            Acids Res. 2023 Nov 30:gkad1125. doi: 10.1093/nar/gkad1125."
COMMENT     Sequence Label: pEJS-HZ08-pAAVsc.U6-tracrRNA
FEATURES             Location/Qualifiers
     source          1..3635
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    135..156
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(171..176,177..194,195..201)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    209..225
                     /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)."
     primer_bind     214..236
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     233..249
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     233..249
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        429..677
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     429..449
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     600..619
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(803..822)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    868..923
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   971..1100
                     /label=AAV2 ITR
     primer_bind     complement(1119..1136)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1119..1135)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1128..1150)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1277..1732
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1364..1383)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1574..1595
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1758..1862
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1863..1931,1932..2723)
                     /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(2081..2100)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2894..3482
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3383..3402
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg
       61 cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt
      121 gagcgcaacg caattaatgt gagttagctc actcattagg caccccaggc tttacacttt
      181 atgcttccgg ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac
      241 agctatgacc atgattacgc cagatttaat taaggcctta attaggctgc gcgctcgctc
      301 gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc
      361 agtgagcgag cgagcgcgca gagagggagt gtagccatgc tctaggaaga tcaattcaat
      421 tcacgcgtga gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg
      481 ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt
      541 gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg
      601 actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt
      661 ggaaaggacg aaacaccgag catagcaagt taaaataagg ctagtccgtt atcaacttga
      721 aaaagtggca ccgagtcggt gcttttttac gcgtcgaccc gggcggcctc gaggacgggg
      781 tgaactacgc ctgaggatcc gatctttttc cctctgccaa aaattatggg gacatcatga
      841 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
      901 gttggaattt tttgtgtctc tcactcggcc taggtagata agtagcatgg cgggttaatc
      961 attaactaca aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg
     1021 ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca
     1081 gtgagcgagc gagcgcgcag ccttaattaa cctaattcac tggccgtcgt tttacaacgt
     1141 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     1201 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     1261 ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     1321 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     1381 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     1441 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     1501 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     1561 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     1621 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     1681 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
     1741 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     1801 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     1861 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     1921 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     1981 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     2041 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     2101 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2161 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     2221 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     2281 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     2341 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     2401 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     2461 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     2521 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     2581 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     2641 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     2701 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     2761 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2821 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     2881 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     2941 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3001 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     3061 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3121 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3181 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3241 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3301 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3361 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3421 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3481 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     3541 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     3601 ctgataccgc tcgccgcagc cgaacgaccg agcgc
//
