LOCUS       sequence-163589-         881 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  AAV backbone with a full length U6/TO promoter driving expression of
            a sgRNA that targets exon 3 of the mouse Tet2 locus. TracrRNA 
            compatible with SpCas9. gRNA transcription is doxycycline 
            dependent..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 881)
  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 881)
  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: 
            pJEP11-AAV-U6/TO-sgRNA(Tet2)-CMV-TetR-P2A-eGFP-KASH-pA
FEATURES             Location/Qualifiers
     source          1..881
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             79..132
                     /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(133..135,136..846)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(133..135,136..846)
                     /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(175..196)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(436..455)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     783..804
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 ttcggccttg aattgatcat atgcggatta gaaaaacaac ttaaatgtga aagtgggtcc
       61 gcggctagcg gcagtggaga gggcagagga agtctgctaa catgcggtga cgtcgaggag
      121 aatcctggcc cagtgagcaa gggcgaggag ctgttcaccg gggtggtacc catcctggtc
      181 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat
      241 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc
      301 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac
      361 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc
      421 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc
      481 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc
      541 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag
      601 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg
      661 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc
      721 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat
      781 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg
      841 tacaagtccg gactcagatc tcgagaggag gaggaggaga c
//
