RAB11A-GFP HDRT Source (pTR 143) vector (V000478)

Basic Vector Information

      • Vector Name:
      • RAB11A-GFP HDRT Source (pTR 143)
      • Antibiotic Resistance:
      • Ampicillin
      • Length:
      • 5814 bp
      • Type:
      • CRISPR, Synthetic Biology
      • Copy Number:
      • High Copy
      • Cloning Method:
      • Gibson Cloning

RAB11A-GFP HDRT Source (pTR 143) vector Vector Map

RAB11A-GFP HDRT Source (pTR 143)5814 bp60012001800240030003600420048005400Amp-RAmpR promoterpBRforEcopGEX 3'pRS-markerIn lacZ geneVN155(I152L)EGFP-CM13 revlac operatorlac promoterCAP binding siteL4440oriAmpR

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 ℃.

RAB11A-GFP HDRT Source (pTR 143) vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                5814 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  DNA sequence source for amplifying an HDR template to tag endogenous
            human RAB11A gene with GFP.
ACCESSION   .
VERSION     .
KEYWORDS    RAB11A-GFP HDRT Source (pTR 143)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5814)
  AUTHORS   Roth TL, Puig-Saus C, Yu R, Shifrut E, Carnevale J, Li PJ, Hiatt J, 
            Saco J, Krystofinski P, Li H, Tobin V, Nguyen DN, Lee MR, Putnam AL,
            Ferris AL, Chen JW, Schickel JN, Pellerin L, Carmody D, 
            Alkorta-Aranburu G, Del Gaudio D, Matsumoto H, Morell M, Mao Y, Cho 
            M, Quadros RM, Gurumurthy CB, Smith B, Haugwitz M, Hughes SH, 
            Weissman JS, Schumann K, Esensten JH, May AP, Ashworth A, Kupfer GM,
            Greeley SAW, Bacchetta R, Meffre E, Roncarolo MG, Romberg N, Herold 
            KC, Ribas A, Leonetti MD, Marson A
  TITLE     Reprogramming human T cell function and specificity with non-viral 
            genome targeting.
  JOURNAL   Nature. 2018 Jul 11. pii: 10.1038/s41586-018-0326-5. doi: 
            10.1038/s41586-018-0326-5.
  PUBMED    29995861
REFERENCE   2  (bases 1 to 5814)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nature. 
            2018 Jul 11. pii: 10.1038/s41586-018-0326-5. doi: 
            10.1038/s41586-018-0326-5."
FEATURES             Location/Qualifiers
     source          1..5814
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     235..254
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(473..577)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     645..663
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(701..723)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     823..842
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1036..1058
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1050..1067
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1051..1067
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2309..2767
                     /codon_start=1
                     /product="improved N-terminal fragment of mVenus for use in
                     bimolecular fluorescence complementation (BiFC) (Kodama and
                     Hu, 2010)"
                     /label=VN155(I152L)
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITA"
     primer_bind     complement(2348..2369)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     2956..2977
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4265..4281)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4265..4281)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4278..4300)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4289..4305
                     /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(4313..4343)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4358..4379
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4496..4513)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4667..5255)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4747..4766)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5426..5814,1..472))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"

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