...difficult-to-manipulate primary cells, achieving knockdown within 1-2 hr of reagent delivery. TRIM-Away is suitable for ...factor 1 controls expression of transcription factor 2, which directly controls expression of a reporter ...modulating the induction of transcription factors 1 and 2. See Aranda-Díaz et al. for more information. The ...Podcast! TRIM-Away: Targeted endogenous protein degradation Article contributed by Mary Gearing Listen to...tool existed for rapid, customizable protein degradation. Enter TRIM-Away - an antibody-based system developed...the protein of interest led to rapid, targeted degradation that is dependent on proteasome function. TRIM-away...

...target sites for editing. PAM sequences can be from 2-6 base pairs in length and follow the DNA sequence... Eric Perkins Mammalian genomic DNA is not just a 2-D linear molecule snaking through our cells’ nuclei... for degradation using AID Article contributed by Alyssa Cecchetelli Auxin Inducible Degradation (AID)...interest fused to a degron, a domain that induces degradation. In plants, auxin binds to TIR1 of the SCF-TIR1... with the degron sequence and ultimately its degradation. While other eukaryotes don’t naturally have ...TIR1 inhibitor, sufficiently suppresses basal degradation of tagged proteins, a common problem of the AID...

...Casilio modules for functional genomics (see Figure 2). We hope that Casilio will aid researchers in a variety...

... efficacy in vivo.  ADAs and Adverse Immune Reactions Anti-drug antibodies (ADAs) are formed when the...foreign antigen, triggering a humoral immune response. ADAs can inactivate the drug and increase drug clearance...various components of the immune system. Figure 2: In-vitro models for immunogenicity prediction of ...drug reaction. This system would somewhat act like a 2-step verification. All approaches have their own limitations...Future Enhancements. Frontiers in drug discovery, 2, 952326. https://doi.org/10.3389/fddsv.2022.952326Joubert...

.... Find Plasmids from the publication at Addgene 2. Green, Alexander A., et al. "Toehold switches: de-novo-designed...expected in patient samples (e.g.: blood, urine). Adapting toehold switches for Zika detection We and other...

...page: SARS-CoV-2 Plasmids : Plasmids that are available or coming soon containing SARS-CoV-2 sequences. SARS-CoV...research. Plasmids SARS-CoV-2 Plasmids Many plasmids containing SARS-CoV-2 sequences are now available...packaging plasmids. SARS-CoV-2 Lentiviral preps now available for several SARS-CoV-2 constructs from the Krogan...SARS-CoV-2 plasmids are curated on a separate Ginkgo Bioworks Plasmid Collection page. SARS-CoV-2 Pooled...Converting Enzyme 2) is the host cell receptor mediating the entry of SARS-CoV and SARS-CoV-2 viruses. ...that primes the SARS-CoV-2 S protein and is involved in virus entry into cells. ( 2 ) FURIN - an enzyme that...of SARS-CoV-2: A Scalable, Easy-to-Deploy, Protocol for Cas13-Based Detection of SARS-CoV-2 Genetic Material...

...immunoglobulin heavy diversity 2-15 D2, IGHD215 IGHD2-2 immunoglobulin heavy diversity 2-2 IGHD22 IGHD2-21 immunoglobulin...heat shock 70kDa protein 2 HSP70-2, HSP70-3 HSPA4 heat shock 70kDa protein 4 APG-2, HS24/P52, MGC131852, ...variable 2-33 (non-functional) IGLV233, V1-9 IGLV2-8 immunoglobulin lambda variable 2-8 IGLV28, V1-2 IGLV3...beta 4 DEFB-2, DEFB102, DEFB2, HBD-2, SAP1 EDN1 endothelin 1 ET1, HDLCQ7 EDN2 endothelin 2 ET2, PPET2 ...receptor subfamily 2, group C, member 1 TR2 NR2C2 nuclear receptor subfamily 2, group C, member 2 TAK1, TR2R1...suppression of tumorigenicity 2 - STC1 stanniocalcin 1 STC STC2 stanniocalcin 2 STC-2, STCRP TAC1 tachykinin,...immunoglobulin heavy diversity 2-21 IGHD221 IGHD2-8 immunoglobulin heavy diversity 2-8 DLR1, IGHD28 IGHD3-10 ...

...of basal H 2 O 2 levels with peroxiredoxin-based probes Real-time monitoring of basal H 2 O 2 levels with...Belousov Hydrogen Peroxide (H 2 O 2 ) Cytosolic or mitochondrial sensor for H 2 O 2 oxidation (roGFP2-Orp1) ...peroxiredoxin-2-based probe. Nat Commun. 2018 Aug 7;9(1):3145. Hadley Sikes Hydrogen Peroxide (H 2 O 2 ) Monitoring...Nat Commun. 2020 Aug 4;11(1):3881 Adam Cohen Hydrogen Peroxide (H 2 O 2 ) Hydrogen peroxide biosensor HyPer7...Calcium Red fluorescent calcium sensors (fRCaMP1/2, GECO1/2) The kinetic mechanisms of fast-decay red-fluorescent...intracellular Zn(2+) homeostasis. Nat Methods. 2009 Oct;6(10):737-40. Maarten Merkx Zinc eZinCh-2 Zn2+ FRET ... eZinCh-2: A Versatile, Genetically Encoded FRET Sensor for Cytosolic and Intraorganelle Zn(2+) Imaging...

...3rd 1, 2, or 4 49,766 arrays Broad GPP kinome Brunello 75314, 75315 (1 plasmid) 75312, 75313 (2 plasmid...GPP genome-wide Brunello 73179 (1 plasmid) 73178 (2 plasmid) Knockout Human Doench and Root 3rd 4 76,441...Broad GPP genome-wide Brie 73632 (1 plasmid) 73633 (2 plasmid) Knockout Mouse Doench and Root 3rd 4 78,637...172651 (Set D) Knockout Human Doench and Root 2nd 2 40,710 Inzolia Human CRISPR/Cas12a Multiplex Knockout...3,052 Broad GPP kinome Brie 75317 (1 plasmid) 75316 (2 plasmid) Knockout Mouse Doench and Root 3rd 4 2,852...MinLibCas9 Library 164896 Knockout Human Garnett 3rd 2 37,722 Green monkey (Chlorocebus sabaeus) sgRNA library...

...FHF2 Human Mouse IgG2a 114527 Anti-JIP-2/IB-2 [N135/37.2R] JIP-2/IB-2 Mouse Mouse IgG2a 114529 Anti-Pan-SAPAP...Anti-CDY1/2/L1/L2 [N143/36R] CDY1/2/L1/L2 Human Mouse IgG2a 206535 Anti-CDY1/2 [N143/38R] CDY1/2 Human Mouse...206763 Neuroligin-2 scFv [L107/90] L107/90 scFv Neuroligin-2 Mouse Mouse 206764 Neuroligin-2 scFv [L107/95...receptor Rat Mouse IgG2a 114484 Anti-Stonin-2 [N346/9R] Stonin-2 Human Mouse IgG2a 114485 Anti-NSD3 [N348...Pan-FHF-A Human Mouse IgG2a 114510 Anti-REEP1/2 [N326D/29R] REEP1/2 Mouse Mouse IgG2a 114511 Anti-Kv3.1b K+ ...Mouse Mouse IgG2a 114547 Anti-Mitofusin-2 [N171/17.4R] Mitofusin-2 Mouse Mouse IgG2a 114548 Anti-Mortalin...Mouse IgG2a 140065 Anti-SCG10/Stathmin-2 [L5/1R] SCG10/Stathmin-2 Rat Mouse IgG2a 140066 Anti-Co-Rest/RCOR1...

...Dilute oligos 1:10 in ddH 2 O ( e.g., 1.0 μl annealed oligos + 9.0 μl ddH 2 O to yield a concentration...been included. Ensure there are 2 x 10 6 cells per CRISPR pair. Resuspend 2 x 10 6 cells in 100 μl of electroporation...), 50-100 ng gDNA, and H 2 O up to 20 μl. Use the primers designed in step 2 above. Conduct PCR for “non-deletion...frequency of indels formed on non-deleted alleles 2 , which are likely to cause frameshift mutations leading...of the Pim1 sgRNA from Table 1 are found in Table 2 . Obtain 24- or 25-mer oligos for each guide and its...sgRNA cleavage sites (“deletion band”; Figures 1 - 2 ). In the absence of deletion, the “deletion band”...Resuspend oligos at a concentration of 100 μM in ddH 2 O. Prepare a 10 μl reaction mix for each guide and...

...111707 Mode #2 Library (NEDD4 WW2 domain) Pooled Library 111708 Mode #2 Library (NEDD4-2 WW2 domain) References...Pooled Library 111705 Mode #2 Library (14-3-3β) Pooled Library 111706 Mode #2 Library (14-3-3σ) Pooled Library...phosphorylation-dependent protein-peptide interactions (“mode #2” expression, Barber et al, Nat. Biotech. 2018). The...fluorescence-activated cell sorting (FACS). The mode #2 phosphosite library with four different phosphobinding... WW2 domain of NEDD4, and the WW2 domain of NEDD4-2; 111705-8 ) is available from Addgene. Screening kinases...iSPI_pSer_Subpool#1 Pooled Library 188527 iSPI_pSer_Subpool#2 Pooled Library 188528 iSPI_pSer_Subpool#3 Pooled Library...pos ctrl Plasmid 111883 pNAS1b split mCherry NEDD4-2 WW2 Plasmid 111882 pNAS1b split mCherry NEDD4 WW2 ...

...par with ZF arrays, if not slightly lower. Figure 2: Simplified representation of the Voytas/Bogdanove...variety of backbones in just a few steps ( Figure 2 ). The Bogdanove group also hosts web-based software...mammalian transcription. Nat Biotechnol. 2011 Feb;29(2):149-53. PMID: 21248753 . TALEs for the masses. Rusk...the Martin-Luther-University Halle-Wittenberg and Adam Bogdanove at Iowa State University published the...by the zinc finger community ( see interview with Adam Bogdanove ), progress on TALe research has been ...-4. Pubmed PMID: 21822235 . See the Q&A with Dr. Adam Bogdanove and his thoughts on the TALe technology...

...transcribed to make the pre-CRISPR RNA (pre-crRNA). (2) The pre-crRNA is processed into individual crRNAs...2015. Class 1 (Multi-subunit effector complex) Class 2 (Single multi-domain effector) Type I (Cas3) Type ... with unprecedented speed and specificity. Figure 2: An overview of CRISPR and NHEJ/HDR. The Cas9/gRNA...regulation of transcription in eukaryotes. Cell . 154(2):442-51. PMID: 23849981 Ishino Y, Shinagawa H, Makino...Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. Cell . 163(3):759-71. PMID: 26422227...finger nucleases (ZFNs) and TALENs. CRISPR: An Adaptive Immune System CRISPR (Clustered Regularly Interspaced...hypothesized that prokaryotes used CRISPR as part of an adaptive immune system - utilizing various CRISPR-associated...

...guide RNA (gRNA) for biochemical purification (Fig. 2). For additional information and protocols, check ... web page . Figure 1: iChIP general scheme Figure 2: enChIP general scheme Fujii Lab Plasmids Individual... of interest 85586 gRNA Cloning Vector Bbs I ver. 2 An empty sgRNA expression vector. sgRNA sequence can... into BbsI site. gRNA scaffold has efficient ver. 2 structure. 92220 pLenti_dCas9-2xAM Lentiviral plasmid...Mohlin S. Biochem Biophys Res Commun. 2018 May 5;499(2):291-298. doi: 10.1016/j.bbrc.2018.03.150. PubMed ...immunoprecipitation (enChIP). Fujita T, Asano Y, Ohtsuka J, Takada Y, Saito K, Ohki R, Fujii H. Sci Rep. 2013 Nov ...

...TGGTGGCGCAGCAGAGGGagctgatccaggaccaGGCCACCGTGAACATCA DLK1 (site #2) TAL3422 & TAL3423 TGAGGTACAGGCAGCTGGtggcgcagcagagggagcTGATCCAGGACCAGGCCA...TGGAAGACCCGTTGGAGAaagagatcccaccaacTGAAATAAAAGATTCAGA hemogen (site #2) TAL3276 & TAL3277 TGATTTGTTTGTTTGCTaggaggaattcggcggCGACTCAGAGACAGAGA...TGTTTCAGCAGAGCCCCGctgaagagctccccatGGAGATGGAAGGAGTGGA hif1al (site #2) TAL3280 & TAL3281 TGCCCTCAGGACTTCTGCacgcctgaactccgcaAGCTTCTGTCTCCAATA...TGGAGCAGGGTGCGAGTGtgtctgagctgaaggaGGCGGTGGGTCGTTTACA park2 (site #2) TAL3334 & TAL3335 TGATCGTTTTCGTGCGGTttaattccagccatggGTTCCCGGTGGAGTTGGA...TGTTAAGGCTCTGTATAGcagtgtgtgtcctggcCACTTGCTGGGCTCAGGA retinitis pigmentosa 2 (X-linked recessive) TAL3526 & TAL3527 TCTTTTTGTGCTGCGCCAcccagcccataatcgaGTCTTCTACAGGCATGAA...TGGAAACCGAGCTGAAGCccccggcgccccagcccaACACCGGGGGCACGGGGA Sox2 (site #2) TAL3368 & TAL3369 TGGTGGGGTAGACTTTCGagaaaatcggtttaaaTGTATAACATGATGGAAA...TGTCCTCATCCCCGTCCCgccgcggcgccaccgtCAGCTTCCACAACATCAA adam15 TAL3016 & TAL3017 TGTTTCTGGCGGACTCTCgtcaccattcttctgtTTGAAGGCACACCTTTGA...

...Regulation Ben Deverman 2:00 PM Cre-DOG and Combinatorial Strategies Connie Cepko 2:45 PM Break 3:00 PM (... Laboratory Gowan Tervo Janelia Research Campus Sarada Viswanathan Janelia Research Campus Kenton Woodard...10:15 AM AAV Retrograde Tracing Gowan Tervo and Sarada Viswanathan 11:00 AM Circuit Tracing Ali Cetin ..., Cold Spring Harbor Laboratory Gowan Tervo and Sarada Viswanathan, Janelia Research Campus Kenton Woodard...

...Sequences Glossary Publications CRISPR Overview Class 2 C lustered R egularly I nterspaced S hort P alindromic...systems enable researchers to target anywhere from 2 to 7 genetic loci by cloning multiple gRNAs into a... is included on the gRNA-containing plasmid, or a 2-plasmid system in which Cas9 must be delivered separately...library (panel E ). Remember - if you are using a 2-vector system, you will need to transduce cells that...Cas enzymes? CasPEDIA is an encyclopedia of Class 2 CRISPR systems with wiki entries describing enzyme...either (1) a lack of gRNA and/or Cas9 expression or (2) a lack of efficient target cleavage in cells expressing...off-target effects by using a single Cas9 nickase and 2 different gRNAs, which bind in close proximity on ...

...8661 p4455 FLAG-hPLIC-2 UBQLN2 Flag CMV ALS Peter Howley 8662 p4456 FLAG-hPLIC-2 NTF UBQLN2 Flag CMV ALS...196208 GB1-A11(2-196)-Strep ANXA11 His, Strep, Tev T7 ALS Lalit Deshmukh 196209 GB1-A11(2-196, G38R)-Strep...196210 GB1-A11(2-196, D40G)-Strep ANXA11 His, Strep, Tev T7 ALS Lalit Deshmukh 196211 GB1-A11(2-196, G175R...196212 GB1-A11(2-196, G189E)-Strep ANXA11 His, Strep, Tev T7 ALS Lalit Deshmukh 196213 GB1-A11(2-52)-Strep ...TLS 1: hTLS.pBSKS(+) FUS T7 ALS David Ron 21828 TLS 2: hTLS.pCDNA1 FUS CMV ALS David Ron 21829 TLS 3: TLS.pRSET.C...ApoE Alzheimer's Sohail Tavazoie 51436 pGL3Basic-ME.2/ApoEpromoter APOE ApoE Alzheimer's Sohail Tavazoie...T7 ALS Nicolas Fawzi 127195 RP1B FUS 1-163 QQ4xSS #2 FUS His T7 ALS Nicolas Fawzi 127196 pJ411 FUS 1-163...

...Cas enzymes? CasPEDIA is an encyclopedia of Class 2 CRISPR systems with wiki entries describing enzyme...Escherichia coli TadA, a tRNA adenine deaminase. Like cytosine base editors, the evolved TadA domain is fused...