LOCUS       sequence-135015-        1099 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses gRNA from hybrid human U6/T7 promoter for both cellular 
            expression and in vitro transcription from same promoter (2 Gs at 
            initiation).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1099)
  AUTHORS   Romanienko PJ, Giacalone J, Ingenito J, Wang Y, Isaka M, Johnson T, 
            You Y, Mark WH
  TITLE     A Vector with a Single Promoter for In Vitro Transcription and 
            Mammalian Cell Expression of CRISPR gRNAs.
  JOURNAL   PLoS One. 2016 Feb 5;11(2):e0148362. doi: 
            10.1371/journal.pone.0148362. eCollection 2016.
  PUBMED    26849369
REFERENCE   2  (bases 1 to 1099)
  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: "PLoS One.";
            date: "2016-02-5"; volume: "11(2):e0148362. doi"; pages: " 
            10.1371/journal.pone.0148362. eCollection 2016"
COMMENT     Sequence Label: pU6T7
FEATURES             Location/Qualifiers
     source          1..1099
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     216..233
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    350..371
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(386..391,392..409,410..416)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    424..440
                     /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     429..451
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     448..464
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     448..464
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     misc_RNA        complement(539..614)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(636..654)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(696..715)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(866..886)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(946..968)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
       61 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      121 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
      181 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
      241 aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
      301 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
      361 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
      421 gaattgtgag cggataacaa tttcacacag gaaacagcta tgaccatgat tacaccaagc
      481 ttgcatgcag gcctctgcag tcgacgggcc cgggatccga tatctagatt taaaaaaagc
      541 accgactcgg tgccactttt tcaagttgat aacggactag ccttatttta acttgctatt
      601 tctagctcta aaactgagac cctctctcgg tctctcctat agtgagtcgt attagcaaga
      661 tatataaagc caagaaatcg aaatactttc aagttacggt aagcatatga tagtccattt
      721 taaaacataa ttttaaaact gcaaactacc caagaaatta ttactttcta cgtcacgtat
      781 tttgtactaa tatctttgtg tttacagtca aattaattct aattatctct ctaacagcct
      841 tgtatcgtat atgcaaatat gaaggaatca tgggaaatag gccctcttcc tgcccgacct
      901 ttctagatgt attcgcgagg taccgagctc gaattctctg gccgtcgttt tacaacgtcg
      961 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     1021 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     1081 gaatggcgaa tggcgcctg
//
