AAV-U6gRNA1-U6gRNA2-TnT-Cre vector (V007261)

Basic Vector Information

      • Vector Name:
      • AAV-U6gRNA1-U6gRNA2-TnT-Cre
      • Antibiotic Resistance:
      • Ampicillin
      • Length:
      • 5958 bp
      • Type:
      • AAV
      • Promoter:
      • cTnT
      • Cloning Method:
      • Restriction Enzyme
      • 5' Primer:
      • ACGACTCACTATAGGCTAGC
      • 3' Primer:
      • AGGGACTTCGGGCACAATCG

AAV-U6gRNA1-U6gRNA2-TnT-Cre vector Vector Map

AAV-U6gRNA1-U6gRNA2-TnT-Cre5958 bp60012001800240030003600420048005400U6 promotergRNA scaffoldU6 promotergRNA scaffoldchimeric intronT7 promoterSV40 NLShGH poly(A) signalAAV2 ITRf1 oripRS-markerpGEX 3'pBRforEcoAmpR promoterAmpRoriAAV2 ITR (alternate)

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 ℃.

AAV-U6gRNA1-U6gRNA2-TnT-Cre vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                5958 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  AAV vector for U6 driven expression of two gRNAs, and cardiomyocyte 
            specific expression of Cre recombinase..
ACCESSION   .
VERSION     .
KEYWORDS    AAV-U6gRNA1-U6gRNA2-TnT-Cre
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5958)
  AUTHORS   Guo Y, VanDusen NJ, Zhang L, Gu W, Sethi I, Guatimosim S, Ma Q, 
            Jardin BD, Ai Y, Zhang D, Chen B, Guo A, Yuan GC, Song LS, Pu WT
  TITLE     Analysis of Cardiac Myocyte Maturation Using CASAAV, A Platform for 
            Rapid Dissection of Cardiac Myocyte Gene Function In Vivo.
  JOURNAL   Circ Res. 2017 Mar 29. pii: CIRCRESAHA.116.310283. doi: 
            10.1161/CIRCRESAHA.116.310283.
  PUBMED    28356340
REFERENCE   2  (bases 1 to 5958)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Circ Res. 
            2017 Mar 29. pii: CIRCRESAHA.116.310283. doi: 
            10.1161/CIRCRESAHA.116.310283."
FEATURES             Location/Qualifiers
     source          1..5958
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        1..241
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        278..353
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        366..606
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        633..708
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     intron          1260..1392
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1375..1394
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1437..1455
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1467..1487
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2554..3030
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2753..2772)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3070..3210
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3070..3199
                     /label=AAV2 ITR
     rep_origin      3285..3740
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3372..3391)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3582..3603
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3757..3776)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3876..3898
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3936..3954)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4022..4126
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4127..4987
                     /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"
     primer_bind     complement(4345..4364)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5158..5746
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5647..5666
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   5808..5937
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"

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