LOCUS       sequence-57101-e         941 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses a guide RNA (gRNA) to target DNMT3b for genome 
            engineering.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 941)
  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 941)
  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_DNMT3b
FEATURES             Location/Qualifiers
     source          1..941
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     protein_bind    52..76
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_RNA        complement(88..163)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(184..432)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(242..261)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(412..432)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     promoter        complement(581..599)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(582..599)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(617..633)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(617..633)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(630..652)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    641..657
                     /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(665..671,672..689,690..695))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    710..731
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(848..865)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gctcgagcgg ccgccagtgt gatggatatc tgcagaattc gccctttaat gccaactttg
       61 tacaagaaag ctgggtctag aaaaaaagca ccgactcggt gccacttttt caagttgata
      121 acggactagc cttattttaa cttgctattt ctagctctaa aacccttggg gcgtgagtaa
      181 ttcggtgttt cgtcctttcc acaagatata taaagccaag aaatcgaaat actttcaagt
      241 tacggtaagc atatgatagt ccattttaaa acataatttt aaaactgcaa actacccaag
      301 aaattattac tttctacgtc acgtattttg tactaatatc tttgtgttta cagtcaaatt
      361 aattctaatt atctctctaa cagccttgta tcgtatatgc aaatatgaag gaatcatggg
      421 aaataggccc tcttcctgcc cgaccttggt accggatcca gtcgactgaa ttggttcctt
      481 taaagcctgc ttttttgtac aaagggcgaa ttccagcaca ctggcggccg ttactagtgg
      541 atccgagctc ggtaccaagc ttgatgcata gcttgagtat tctatagtgt cacctaaata
      601 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
      661 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
      721 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
      781 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
      841 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
      901 ctcactcaaa ggcggtaata cggttatcca cagaatcagg g
//
