1982-2-1 · pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro was a gift from Feng Zhang (Addgene plasmid # 42229) Multiplex Genome Engineering Using CRISPR/Cas Systems. Cong L Ran FA Cox D Lin S Barretto R Habib N Hsu
pX260 pX330 pX330A-1x2 pX330S-2 pX334 pX335 PX551 PX552 pX600 pX601 pX602 PX851 PX852 PX853 PX854 PX855 PX856 pY010 (pcDNA3.1-hAsCpf1) pY016 (pcDNA3.1-hLbCpf1) sgRNA(MS2) cloning backbone SP-Cas9 SP-dCas9-VPR VP12
2015-6-5 · Figure S1. CRISPR-mediated targeting of the OCT4 locus through drug selection. (A) The vector map of the px260 plasmid for expressing Cas9 and crRNA/tracrRNA. (B) Four crRNAs were designed to target the stop codon of the OCT4 locus.Each crRNA was cloned into the px260 vector and transfected into 293T cells.
2019-9-2 · PX260 This plasmid separately encodes a human codon-optimized SpCas9 a tracrRNA and customizable crRNA. #42230 PX330 A human codon-optimized SpCas9 and chimeric guide RNA expression plasmid. #48138 PX458 GFP Cas9 from S. pyogenes
1982-2-1 · pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro was a gift from Feng Zhang (Addgene plasmid # 42229) Multiplex Genome Engineering Using CRISPR/Cas Systems. Cong L Ran FA Cox D Lin S Barretto R Habib N Hsu
2021-4-27 · 2.5 mg pX260 plasmid and 2.5 mg donor plasmid with GFP by electroporator (Nepa Gene Chiba Japan). Transfected single cells were recovered in NutriStem hPSC XF Medium (Biological Industries Beit-Haemek Israel) and seeded on Matrigel-coated plates. Four days after electroporation the culture medium was changed back to mTeSR1 medium and
2020-7-7 · PX260 This plasmid separately encodes a human codon-optimized SpCas9 a tracrRNA and customizable crRNA. #42230 PX330 A human codon-optimized SpCas9 and chimeric guide RNA expression plasmid. #48138 PX458 GFP Cas9 from S. pyogenes
2021-7-2 · The first delivery pattern is a plasmid such as pX260 and pX335 which encodes both sgRNA and Cas9. The second delivery pattern is a mixture of sgRNA and Cas9 mRNA. The third delivery pattern is Cas9/sgRNA ribonucleoprotein (RNP). 28 29 Various advantages and shortcomings lie in each delivery pattern for clinical translation.
2017-11-28 · The pX260 system also known as the pX334 system contains three cassettes including a CRISPR RNA array tracrRNA and S. pyogenes Cas9 (or the Cas9 D10A nickase). The plasmid is digested with a restriction enzyme and then ligated with an annealed oligonucleotide that is designed for a specific targeting site.
2020-12-14 · 1. Digest 1ug of plasmid with . Bbs. I for. 30 min at 37°C 1 ug Plasmid . 1 ul FastDigest . Bbs. I (Fermentas) 1 ul FastAP (Fermentas) 2 ul 10X FastDigest Buffer . X ul ddH. 2 O 20 ul total 2. Gel purify digested plasmid using. QIAquick Gel Extraction Kit and elute in EB. 3. Phosphorylate and anneal each pair of oligos 1 ul oligo 1 (100uM)
px260. In Category. Educational Resources. Filter By. Expression. Mammalian Expression (1) Vector Types. CRISPR (1) Plasmid Type.
Show Static Map. hCas9 9553 bp. Cas9. 748 .. 4851 = 4104 bp. 1368 amino acids = 158.4 kDa. Product Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system. generates RNA-guided double strand breaks in DNA.
pX260 #42229 pX330 #42230 pX335 #42335 pX458 #48138 pX458M EZ-GuideXH pX459M EZ-GuideXH EZ-GuideXH pX461 #48140 pX552 #60958 pX601 #61591 pX602 #61593 LentiCRISPR V2 #52961 pLen
2016-3-4 · For gRNA expression screening specific reverse primer (pX260-crRNA-3′/R Table I.3 in S1 Experimental Procedures) was used in RT reaction followed by standard PCR using target A or B
2014-7-18 · Plasmid Preparation. DNA segment expressing long-term repeats (LTR)-A or LTR-B for precrRNA was cloned into the pX260 vector that contains the puromycin selection gene (Addgene plasmid #42229). DNA segment expressing LTR-C or LTR-D for the chimeric crRNA-tracrRNA was cloned into the pX330 vector (Addgene plasmid #42230). Both vectors contain
2014-9-25 · Linearize plasmid PX260 (Addgene plasmid #42229) using the NcoI restriction site which is located immediately upstream of the initial codon of the NLS-hSpCas9-NLS expression cassette. Ligate the
2017-11-28 · The pX260 system also known as the pX334 system contains three cassettes including a CRISPR RNA array tracrRNA and S. pyogenes Cas9 (or the Cas9 D10A nickase). The plasmid is digested with a restriction enzyme and then ligated with an annealed oligonucleotide that is designed for a specific targeting site.
2020-12-14 · 1. pX260 (or pX334) S. pyogenes Cas9 (or Cas9 D10A nickase) CRISPR RNA array tracrRNA This plasmid contains three expression cassettes. In order to target a given site the plasmid can be digested using BbsI and a pair of annealed oligos (design is indicated below) can be cloned into the CRISPR array.
pX260. Zhang lab vector for expressing Cas9 a tracrRNA and a guide sequence. Also known as pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro. To see this sequence with restriction sites features and translations please download
2020-7-7 · PX260 This plasmid separately encodes a human codon-optimized SpCas9 a tracrRNA and customizable crRNA. #42230 PX330 A human codon-optimized SpCas9 and chimeric guide RNA expression plasmid. #48138 PX458 GFP Cas9 from S. pyogenes
2014-9-25 · Linearize plasmid PX260 (Addgene plasmid #42229) using the NcoI restriction site which is located immediately upstream of the initial codon of the NLS-hSpCas9-NLS expression cassette. Ligate the
2020-8-14 · px260 (Addgene plasmid no. 42229) pCMV-BE3 (Addgene plasmid no. 73021) pCAG-Cre (Yang lab no. ZP156) FastDigest BbsI (Thermo Fisher Scientific . no. FD1014)
2014-8-7 · Puro-Cas9 donor (Figure S1E) was constructed by replacing EGFP in the TRE-TIGHT-EGFP-BW plasmid (Addgene plasmid 22077) (Hockemeyer et al. 2011) with the human codon-optimized Streptococcus pyogenes Cas9 cDNA amplified by PCR from pX260 (Addgene plasmid
2020-8-14 · px260 (Addgene plasmid no. 42229) pCMV-BE3 (Addgene plasmid no. 73021) pCAG-Cre (Yang lab no. ZP156) FastDigest BbsI (Thermo Fisher Scientific . no. FD1014)
2014-9-25 · Linearize plasmid PX260 (Addgene plasmid #42229) using the NcoI restriction site which is located immediately upstream of the initial codon of the NLS-hSpCas9-NLS expression cassette. Ligate the
2019-7-16 · PX260 This plasmid separately encodes a human codon-optimized SpCas9 a tracrRNA and customizable crRNA. #42230 PX330 A human codon-optimized SpCas9 and chimeric guide RNA expression plasmid. #48138 PX458 GFP Cas9 from S. pyogenes
a plasmid such as pX260 and pX335 which encodes both sgRNA and Cas9. The second delivery pattern is a mixture of Scheme 1 Key considerations in designing CRISPR/Cas9-carrying nanoparticles for therapeutic genome editing. Fig. 1 Delivery patterns of CRISPR/Cas9 components. The first delivery
2020-7-7 · PX260 This plasmid separately encodes a human codon-optimized SpCas9 a tracrRNA and customizable crRNA. #42230 PX330 A human codon-optimized SpCas9 and chimeric guide RNA expression plasmid. #48138 PX458 GFP Cas9 from S. pyogenes
Plasmid pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro from Dr. Feng Zhang s lab contains the insert humanized S. pyogenes Cas9 and is published in Science. 2013 Jan 3. This plasmid is available through Addgene.
Show Static Map. hCas9 9553 bp. Cas9. 748 .. 4851 = 4104 bp. 1368 amino acids = 158.4 kDa. Product Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system. generates RNA-guided double strand breaks in DNA.
In order to target a given site the plasmid can be digested using BbsI and a pair of annealed oligos (design is indicated below) can be cloned into the CRISPR array. The oligos is designed based on the target site sequence (30bp) and needs to be flanked on the 3′ end by a 3bp NGG PAM sequence. Genbank Map of Backbone Plasmid PX260 (rev
2017-1-22 · To construct pSicoR-sgRNAs-mCherry-Cas9 vector the puromycin fragment in pSico-EF1-mCh-Puro (Addgene Plasmid 31845) was replaced with the NLS-Cas9-NLS fragment from pX260 (Addgene Plasmid 42229) .Then the gRNA scaffold with the cloning site containing two BsmBI sites was inserted downstream of the U6 promoter.The schematic of pSicoR-sgRNA-mCherry-Cas9 vector was
pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro was a gift from Feng Zhang (Addgene plasmid # 42229) Multiplex Genome Engineering Using CRISPR/Cas Systems. Cong L Ran FA Cox D Lin S Barretto R Habib N Hsu
Why does thePX260plasmid use 30nt oligos for cloning butPX330only uses 20nt The reason for the 30nt region on the upper is because the top figure shows the construct from our Split RNA design where the pre-processing form of crRNA is expressed and a tracrRNA (the helper RNA that facilitate the processing of the crRNA) is also expressed from a separate promoter.
Plasmid pX330-U6-Chimeric_BB-CBh-hSpCas9 from Dr. Feng Zhang s lab contains the insert humanized S. pyogenes Cas9 and is published in Science. 2013 Jan 3. This plasmid is available through Addgene. More than 20 requests
2016-3-4 · For gRNA expression screening specific reverse primer (pX260-crRNA-3′/R Table I.3 in S1 Experimental Procedures) was used in RT reaction followed by standard PCR using target A or B
2020-6-29 · xCas9(3.7)-ABE(7.10)(Plasmid #108382)HH-CAS-065 xCas9 3.7PAM PAM xCas9(3.7)–BE3
This plasmid is referred to as pX260-MYC 3′ TBE1. The oligonucleotide sequences corresponding to the guide RNA are listed in Table S1. To design the MYC 3′ WRE-specific CRISPR reporter plasmid we employed a stepwise approach using standard PCR and sub-cloning procedures. The pEGFP-N1 plasmid (Clontech 6085-1 Mountain View CA USA
2017-11-28 · The pX260 system also known as the pX334 system contains three cassettes including a CRISPR RNA array tracrRNA and S. pyogenes Cas9 (or the Cas9 D10A nickase). The plasmid is digested with a restriction enzyme and then ligated with an annealed oligonucleotide that is designed for a specific targeting site.
2020-12-14 · 1. pX260 (or pX334) S. pyogenes Cas9 (or Cas9 D10A nickase) CRISPR RNA array tracrRNA This plasmid contains three expression cassettes. In order to target a given site the plasmid can be digested using BbsI and a pair of annealed oligos (design is indicated below) can be cloned into the CRISPR array.
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