dCas9g1pac vector (V010023)

LOCUS       40924_595        9355 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector dCas9g1pac, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9355)
  AUTHORS   McInally SG, Hagen KD, Nosala C, Williams J, Nguyen K, Booker J, 
            Jones K, Dawson SC.
  TITLE     Robust and stable transcriptional repression in Giardia using 
            CRISPRi
  JOURNAL   Mol. Biol. Cell, mbcE18090605 (2018) In press
  PUBMED    30379614
REFERENCE   2  (bases 1 to 9355)
  AUTHORS   Hagen KD, McInally SG, Dawson SC.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-MAR-2018) Dept of Microbiology and Molecular Genetics,
            University of California, Davis, One Shields Ave, Davis, CA 95616, 
            USA
REFERENCE   3  (bases 1 to 9355)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 9355)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol. Biol. 
            Cell, mbcE18090605 (2018) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (08-MAR-2018) Dept of Microbiology and Molecular Genetics, 
            University of California, Davis, One Shields Ave, Davis, CA 95616, 
            USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9355
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      17..605
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    893..914
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        929..959
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    967..983
                     /label=lac operator
                     /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     991..1007
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1025..1043
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     5'UTR           1064..1204
                     /gene="malate dehydrogenase"
     gene            1064..1204
                     /gene="malate dehydrogenase"
                     /label=malate dehydrogenase
     CDS             1205..5308
                     /label=dCas9
                     /note="catalytically dead mutant of the Cas9 endonuclease
                     from the Streptococcus pyogenes Type II CRISPR/Cas system"
     misc_feature    5315..5335
                     /gene="dCas9"
                     /label=SV40 NLS
                     /note="SV40 NLS"
     CDS             5315..5335
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5350..5366)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             5378..5404
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
     CDS             5408..5434
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
     CDS             5438..5464
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
     misc_feature    5489..5590
                     /gene="dCas9"
                     /label=Giardia NLS
                     /note="Giardia NLS"
     3'UTR           5603..5727
                     /gene="alpha tubulin"
     gene            5603..5727
                     /gene="alpha tubulin"
                     /label=alpha tubulin
     5'UTR           5735..6008
                     /gene="gamma giardin"
     gene            5735..6008
                     /gene="gamma giardin"
                     /label=gamma giardin
     CDS             6015..6608
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
     3'UTR           6618..6736
                     /gene="glutamate dehydrogenase"
     gene            6618..6736
                     /gene="glutamate dehydrogenase"
                     /label=glutamate dehydrogenase
     misc_feature    6949..6962
                     /label=region with BbsI sites for cloning
                     /note="region with BbsI sites for cloning"
     misc_RNA        6965..7040
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(7267..7285)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7292..7308)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(7449..7904)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        8236..8340
                     /label=AmpR promoter
     CDS             8341..9198
                     /label=AmpR
                     /note="beta-lactamase"