LOCUS       Exported                3493 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector U6p-gRNA1, complete sequence.
ACCESSION   KJ796490
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3493)
  AUTHORS   Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK.
  TITLE     Multiplexed and programmable regulation of gene networks with an 
            integrated RNA and CRISPR/Cas toolkit in human cells
  JOURNAL   Mol. Cell 54 (4), 698-710 (2014)
  PUBMED    24837679
REFERENCE   2  (bases 1 to 3493)
  AUTHORS   Perli SD.
  TITLE     Direct Submission
  JOURNAL   Submitted (04-MAY-2014) Electrical Engineering and Computer Science,
            Massachusetts Institute of Technology, 77, Massachusetts Avenue, 
            Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 3493)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..3493
                     /organism="Cloning vector U6p-gRNA1"
                     /mol_type="other DNA"
                     /db_xref="taxon:1520553"
     regulatory      33..42
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      37..46
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     protein_bind    61..85
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      90..95
                     /regulatory_class="terminator"
                     /note="PolIII terminator"
     misc_feature    96..191
                     /label=gRNA1
                     /note="gRNA1"
     misc_RNA        complement(97..172)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      193..510
                     /regulatory_class="promoter"
                     /gene="U6"
     gene            193..510
                     /gene="U6"
                     /label=U6
     promoter        complement(201..441)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     promoter        complement(618..636)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(654..670)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    678..694
                     /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(702..732)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    747..768
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(1056..1644)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1815..2675)
                     /codon_start=1
                     /product="ampicillin resistance protein"
                     /label=ampicillin resistance protein
                     /protein_id="AIJ03089.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(2676..2780)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2858..3313)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     3454..3470
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3477..2
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"