... stable Cas expression under low temperature conditions, thereby enabling efficient CRISPR-Cas mediated... our strategy using the pMBP-LbCas12a and pC013-Twinstrep-SUMO-huLwCas13a plasmids from Addgene, which...

...    References and resources References Janeway CA Jr, Travers P, Walport M, et al. Immunobiology: The...https://www.ncbi.nlm.nih.gov/books/NBK27106/  Janeway CA Jr, Travers P, Walport M, et al. Immunobiology: The...complement system proteins, which can initiate a cascade that helps attack extracellular pathogens.  Transportation...encode those proteins are designated by the lower-case Greek letters μ, δ, γ, ε, and α. Some isotypes have...

...advantage to using the CRISPR/Cas system for genome engineering is the fact that Cas9 can be easily programmed...https://doi.org/10.1016/j.cell.2013.08.021 Waters CA, Strande NT, Pryor JM, Strom CN, Mieczkowski P, Burkhalter.... Repair of Cas9-induced breaks by NHEJ As illustrated below, NHEJ-mediated repair of Cas9-generated breaks...discuss NHEJ, and how it impacts what happens to Cas9-mediated DSBs in the genome. Non-homologous end...consequently the principal means by which CRISPR/Cas9-introduced breaks are repaired. The following factors...introduce indels. Given the end structure of the Cas9 DSB (blunt or near-blunt ends without nucleotide...population of cells constitutively expressing a targeted Cas9 should possess indels in the majority of their chromosomes...

...of CRISPR, Addgene gave to me: a miniature CRISPR-Cas system for mammalian use. On the second day of CRISPR...: efficient CRISPR editing with a hypercompact Cas12f1 and engineered guide RNAs. On the sixth day of ...programmed genome editing by a miniature CRISPR-Cas12f nuclease. On the eighth day of CRISPR, Addgene ...the eleventh day of CRISPR, Addgene gave to me: a Cas9 nickase 101 blog post.  On the twelfth day of CRISPR...

...Yang, J. et al. (7) wished to trigger intracellular Ca++ release with an optogenetic receptor, melanopsin...composed of Venus fluorescent protein, the Troponin C Ca++ binding domain, and NanoLuc® Luciferase. CalFlux...stimulation with NanoLuc-based luminescence (BRET) Ca++ sensing. Nature Comm. 7, 13268. PubMed PMID: 27786307...endogenous tagged genes generated through CRISPR/Cas9 knock-in. Finally, NLuc is very inviting for endogenous...of photobleaching, autofluorescence, and, in the case of exciting cyan-excitable donors, phototoxicity...

...Thakore PI, Majoros WH, Reddy TE, Gersbach CA. Multiplex CRISPR/Cas9-based genome editing for correction of...Ran FA, Yan WX, Asokan A, Zhang F, Duan D, Gersbach CA. In vivo genome editing improves muscle function ...Perez-Pinera P, Brown MT, Majoros WH, Reddy TE, Gersbach CA. Correction of dystrophin expression in cells from...long period of time. High fidelity Cas9s such as eSpCas9 and SpCas9-HF should be explored to reduce off-target...mutation in dystrophin exon 23. Long et al. used SpCas9 in conjunction with AAV9, whereas Nelson et al....al. and Tabebordbar et al. used the shorter SaCas9 with AAV8 and AAV9, respectively. Although each study... it’s important to realize that DMD is a simpler case than other genetic diseases we’d like to treat with...

... 11th day of CRISPR, Addgene gave to me: Cas-E-le-ven! (Cas11)🎵 On the 10th day of CRISPR, Addgene gave...🎵 On the 3rd day of CRISPR, Addgene gave to me: dCas9-based gene editing for cleavage-free genomic knock-in...day of CRISPR, Addgene gave to me: High-fidelity Cas13 variants for targeted RNA degradation with minimal...

...al. Chemically modified guide RNAs enhance CRISPR-Cas genome editing in human primary cells. Nat Biotechnol...Photoswitchable gRNAs for Spatiotemporally Controlled CRSIPR-Cas-based Genomic Regulation. ACS 6, 5, 695-703 (2020...surprise then that stability of the RNA used in CRISPR-Cas9 experiments – the “gRNA” – suffers from the same...still effectively serving their function of guiding Cas9 to generate a targeted break. These advances in ...modifications The recognition sequence for CRISPR-Cas9 targeting is led by a ~20 bp gRNA which is part ...trans-activating crRNA (tracrRNA) which interacts with the Cas9 protein. These RNA molecules are vulnerable to attack...partially DNA gRNA is surprisingly well-tolerated by the Cas9 protein and can increase on-target efficiency. In...

... it’s easy to switch out reporters, Cas proteins, guide cassettes, promoters, N’ and/or C’ terminus, and...specific to the type of module it is (Pol 2 protomer, Cas protein, ect...). Whatever individual components ...above selections.) CRISPR enzyme (Cas9 [sp], Cas9 [other], Cas12a, RNA-targeting) CRISPR mechanism ...destination vectors contain an ampicillin resistance cassette and an origin of replication, which are not schematized...using more than six or seven components — in most cases, that's making two or more plasmids — it may be ...

...Hypercompact genome editor CRISPR-Casɸ By Melina Fan The Doudna Lab adds to the CRISPR-Cas toolbox with their discovery...well-characterized Cas enzymes, which rely on multiple active sites. In addition, Casɸ has a minimal PAM...three Casɸ orthologs, Casɸ-1, Casɸ-2, and Casɸ-3. This system is notable for several reasons. Casɸ is only...70kDa, with a molecular weight roughly half of Cas9 or Cas12a. This is in part due to the fact that it only...demonstrate that CRISPR-Casɸ is active in vitro as well as in human and plant cells. Casɸ from Biggiephage...Blue-light inducible Cre for use in bacteria CRISPR-Casɸ mGold, a new yellow fluorescent protein New CRISPR...excision or insertion, inversion, translocation or cassette exchange. By utilizing recombinases to manipulate...