LOCUS       sequence-163802-        1046 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  AAV backbone with a full length H1/TO promoter driving expression of
            a sgRNA. The gRNA can be inserted via Sapl sites. TracrRNA 
            compatible with SpCas9. gRNA transcription is doxycycline 
            dependent..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1046)
  AUTHORS   de Solis CA, Ho A, Holehonnur R, Ploski JE
  TITLE     The Development of a Viral Mediated CRISPR/Cas9 System with 
            Doxycycline Dependent gRNA Expression for Inducible In vitro and In 
            vivo Genome Editing.
  JOURNAL   Front Mol Neurosci. 2016 Aug 18;9:70. doi: 10.3389/fnmol.2016.00070.
            eCollection 2016.
  PUBMED    27587996
REFERENCE   2  (bases 1 to 1046)
  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: "Front Mol 
            Neurosci."; date: "2016-08-18"; volume: "9:70. doi"; pages: " 
            10.3389/fnmol.2016.00070. eCollection 2016"
COMMENT     Sequence Label: 
            pJEP12-AAV-H1/TO-L-sgRNA(Empty)-CMV-TetR-P2A-eGFP-KASH-pA
FEATURES             Location/Qualifiers
     source          1..1046
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             76..129
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(130..132,133..843)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(130..132,133..843)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(172..193)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(433..452)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     780..801
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 ggccttgaat tgatcatatg cggattagaa aaacaactta aatgtgaaag tgggtccgcg
       61 gctagcggca gtggagaggg cagaggaagt ctgctaacat gcggtgacgt cgaggagaat
      121 cctggcccag tgagcaaggg cgaggagctg ttcaccgggg tggtacccat cctggtcgag
      181 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
      241 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
      301 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
      361 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
      421 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
      481 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
      541 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
      601 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
      661 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
      721 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
      781 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
      841 aagtccggac tcagatctcg agaggaggag gaggagacag acagcaggat gccccacctc
      901 gacagccccg gcagctccca gccgagacgc tccttcctct caagggtgat cagggcagcg
      961 ctaccgttgc agctgcttct gctgctgctg ctgctcctgg cctgcctgct acctgcctct
     1021 gaagatgact acagctgcac ccaggc
//
