LOCUS       sequence-67049-e         977 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses a guide RNA (gRNA) to target human AAVS1 (T1 target 
            sequence) for genome engineering.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 977)
  AUTHORS   Mali P, Yang L, Esvelt KM, Aach J, Guell M, Dicarlo JE, Norville JE,
            Church GM
  TITLE     RNA-Guided Human Genome Engineering via Cas9.
  JOURNAL   Science. 2013 Feb 15;339(6121):823-6. doi: 10.1126/science.1232033
  PUBMED    23287722
REFERENCE   2  (bases 1 to 977)
  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: "10.1126/science"; 
            journalName: "Science"; date: "2013-02-15- 15"; volume: "339"; 
            issue: "6121"; pages: "823-6"
COMMENT     Sequence Label: gRNA_AAVS1-T1
FEATURES             Location/Qualifiers
     source          1..977
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        122..370
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     122..142
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     293..312
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        391..466
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    complement(478..502)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        complement(587..605)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(588..605)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(623..639)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(623..639)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(636..658)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    647..663
                     /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(671..677,678..695,696..701))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    716..737
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(854..871)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 atgcatgctc gagcggccgc cagtgtgatg gatatctgca gaattcgccc tttgtacaaa
       61 aaagcaggct ttaaaggaac caattcagtc gactggatcc ggtaccaagg tcgggcagga
      121 agagggccta tttcccatga ttccttcata tttgcatata cgatacaagg ctgttagaga
      181 gataattaga attaatttga ctgtaaacac aaagatatta gtacaaaata cgtgacgtag
      241 aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa tggactatca
      301 tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct tgtggaaagg
      361 acgaaacacc gtcccctcca ccccacagtg gttttagagc tagaaatagc aagttaaaat
      421 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt tttctagacc
      481 cagctttctt gtacaaagtt ggcattaaag ggcgaattcc agcacactgg cggccgttac
      541 tagtggatcc gagctcggta ccaagcttga tgcatagctt gagtattcta tagtgtcacc
      601 taaatagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
      661 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
      721 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
      781 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
      841 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
      901 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
      961 agaacatgtg agcaaaa
//
