P1-EYFP-Triplex-28-gRNA2-28-pA vector (V009797)

Basic Vector Information

      • Vector Name:
      • P1-EYFP-Triplex-28-gRNA2-28-pA
      • Antibiotic Resistance:
      • Ampicillin
      • Length:
      • 4592 bp
      • Type:
      • Cloning vector
      • Source/Author:
      • Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK.

P1-EYFP-Triplex-28-gRNA2-28-pA vector Vector Map

P1-EYFP-Triplex-28-gRNA2-28-pA4592 bp600120018002400300036004200oriampicillin resistance proteinAmpR promoterf1 oripause site8x gRNA1 binding sitesMinimal Ade MLPPhiYFPMALAT1_3 triplexCsy4 rptgRNA2Csy4 rpt

Plasmid Resuspension Protocol:

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.

3. Close the tube and incubate for 10 minutes at room temperature.

4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.

5.Store the plasmid at -20 ℃.

P1-EYFP-Triplex-28-gRNA2-28-pA vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                4592 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector P1-EYFP-Triplex-28-gRNA2-28-pA, complete sequence.
ACCESSION   KJ796501
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4592)
  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 4592)
  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 4592)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..4592
                     /organism="Cloning vector P1-EYFP-Triplex-28-gRNA2-28-pA"
                     /mol_type="other DNA"
                     /db_xref="taxon:1520549"
     rep_origin      complement(99..687)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(858..1718)
                     /codon_start=1
                     /product="ampicillin resistance protein"
                     /label=ampicillin resistance protein
                     /protein_id="AIJ03100.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(1719..1823)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1850..2305
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     misc_feature    2496..2587
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     misc_feature    2601..2962
                     /label=8x gRNA1 binding sites
                     /note="8x gRNA1 binding sites"
     regulatory      2969..3030
                     /regulatory_class="promoter"
                     /note="Minimal Ade MLP"
     CDS             3037..3741
                     /codon_start=1
                     /product="Phialidium yellow fluorescent proteim"
                     /label=PhiYFP
                     /note="mammalian codon-optimized"
                     /translation="MSSGALLFHGKIPYVVEMEGDVDGHTFSIRGKGYGDASVGKVDAQ
                     FICTTGDVPVPWSTLVTTLTYGAQCFAKYGPELKDFYKSCMPDGYVQERTITFEGDGNF
                     KTRAEVTFENGSVYNRVKLNGQGFKKDGHVLGKNLEFNFTPHCLYIWGDQANHGLKSAF
                     KICHEIAGSKGDFIVADHTQMNTPIGGGPVHVPEYHHMSYHVKLSKDVTDHRDNMSLTE
                     TVRAVDCRKTYL"
     gene            3037..3738
                     /gene="EYFP"
                     /label=EYFP
     misc_feature    3751..3860
                     /label=MALAT1_3 triplex
                     /note="MALAT1_3 triplex"
     misc_feature    3911..3938
                     /label=Csy4 rpt
                     /note="Csy4 rpt"
     protein_bind    3916..3930
                     /label=Csy4 site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease 
                     (Haurwitz et al., 2010)"
     misc_feature    3939..4042
                     /label=gRNA2
                     /note="gRNA2"
     misc_RNA        3959..4034
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    4043..4070
                     /label=Csy4 rpt
                     /note="Csy4 rpt"
     protein_bind    4048..4062
                     /label=Csy4 site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease 
                     (Haurwitz et al., 2010)"
     regulatory      4137..4358
                     /regulatory_class="polyA_signal_sequence"
                     /gene="SV40"
     gene            4137..4358
                     /gene="SV40"
                     /label=SV40
     polyA_signal    4146..4267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"

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