...references are below. Pooled genome-wide human SAM library SAM can be combined with a human genome-wide library...please visit the Addgene SAM library page . Targeted gene activation using SAM There are two sets of 3 ...CRISPR Blog Posts Jump to: SpCas9 Plasmids GeCKO SAM AAV - In vivo Delivery AAV - Cas9 Mouse CRISPR ( ...information. Full references are below. Return to top Cas9 SAM transcriptional activation plasmids and screening...library CRISPR/Cas9 Synergistic Activation Mediator (SAM) is an engineered protein complex for the transcriptional...available for mammalian endogenous gene activation using SAM: Addgene plasmids #61422 - #61424 were designed for...sites for insertion of guide sequences Zhang Lab SAM Cloning Protocol 321.5 KB Return to top AAV - In vivo...

...70,290 SAM v1 - 3 plasmid system 1000000075 (Puromycin) Activation Mouse Zhang 3rd 3 69,716 SAM v2 - 2...3rd 1, 2, or 4 49,766 arrays Broad GPP kinome Brunello 75314, 75315 (1 plasmid) 75312, 75313 (2 plasmid... Library 141438 Knockout Human Yeo 3rd 10 12,472 SAM v1 - 3 plasmid system 1000000057 (Zeocin) 1000000074...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...

... has increased stability and light output Sanjiv Sam Gambhir 104587 pHIV-iRFP720-E2A-Luc Firefly EF1α ...fusing NanoLuc® and Venus fluorophore to Troponin C Ca 2+ binding domain. Nano-lanterns , Enhanced Nano-lanterns...transfection Feng Zhang 21471 pLenti PGK V5-LUC Neo (w623-2) Firefly PGK Lentiviral expression of firefly luciferase...

...different activators exist, including the multi-plasmid SAM system. Select Expression System To use CRISPR, you...either (1) a lack of gRNA and/or Cas9 expression or (2) a lack of efficient target cleavage in cells expressing...efficacious than gRNAs containing a C nucleotide at the same position in spite of being a perfect match for the...

...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...Run samples on 2% agarose gel at 10 V/cm using 1x Tris-acetate-EDTA (TAE) buffer. Examine samples for .... Pick 2 - 3 colonies and inoculate into a mini-prep culture. Perform mini-prep for each sample and sequence...deletion. Run sample in thermocycler and run the following program: 65 °C for 6 min, 98 °C for 2 min to extract... gDNA from clones. Run sample in thermocycler: 65 °C for 6 min and 98 °C for 2 min to extract gDNA. Screen...

...fusion none 2, 5 Richie 121539 pAAV SYN1 HA-hM3D(Gq) hM3D(Gq) - Activation HA fusion none 2, 5 Richie 50467...none 2, 8 Roth 50476 pAAV-CaMKIIa-hM3D(Gq)-mCherry hM3D(Gq) - Activation mCherry fusion none 1, 2, 5, ...(Gq) - Activation mCherry fusion Cre-dependent 1, 2, 5, 8, 9, rg*, PHPeB Roth 44362 pAAV-hSyn-DIO-hM4D...(Gi) - Inhibition mCherry fusion Cre-dependent 1, 2, 5, 8, 9, rg*, PHPeB Roth 65417 pAAV-hSyn-dF-HA-KORD-IRES-mCitrine...mCherry hM3D(Gq) - Activation mCherry fusion none 1, 2, 5, 8, 9, rg* Roth 50475 pAAV-hSyn-hM4D(Gi)-mCherry...mCherry hM4D(Gi) - Inhibition mCherry fusion none 1, 2, 5, 8, 9, rg* Roth 52536 rAAV-CAG::FLEX-rev:: hM4D-...mCherry hM4D(Gi) - Inhibition mCherry fusion none 1, 2, 5, 8, 9 Roth 75033 pAAV CD68-hM4D(Gi)-mCherry hM4D...

...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...encoded Ca(2+) indicator with enhanced two-photon absorption. Neurophotonics. 2024 Apr;11(2):024207. doi... 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...

...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,...FLJ99988, FLT2, HBGFR, KAL2, N-SAM, OGD FGFR2 fibroblast growth factor receptor 2 BEK, BFR-1, CD332, CEK3, ...

...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 ...

...suggests that titers obtained using ddPCR are generally 2–3 fold higher than those achieved using the standard...catalog item (see maps section for images). Figure 2: AAV Pro cells were transduced with either pAAV-Ef1a-mCherry-IRES-Cre... a new vial of early passage cells. Approximately 2 weeks post-thaw, cell culture supernatant is tested...of the viral capsid proteins VP1, VP2 and VP3. Samples of viral preparations are subjected to polyacrylamide...proteins as a fraction of total protein present in the sample is also determined and used to determine purity...shown relative to the total protein present in the sample. Sterility Viral vector preparations are added ...rigorous barcode matching system that follows the sample from the DNA tube all the way to the cleared viral...

... mutations in the gene methyl-CpG binding protein 2 ( MECP2 ). MECP2 Rett syndrome is an X-linked disorder...X-linked MECP2, encoding methyl-CpG-binding protein 2. Nat Genet . 23, 185–188. (Link opens in a new window... Cuddapah et al. 2014. Methyl-CpG-binding protein 2 (MECP2) mutation type is associated with disease severity... Specific mutations in methyl-CpG-binding protein 2 confer different severity in Rett syndrome. Neurology...males with mutations in Methyl-CpG binding protein 2. Am J Med Genet B Neuropsychiatr Genet . 180, 55–67...variability in symptoms even within patients with the same mutation, the location and type of MECP2 mutation...cell line repository (Link opens in a new window) Sampled - one of the world's largest biorepositories (Link...

...overview of the most common methods. BioBrick BioBrick-2 BglBrick Silver Standard Freiburg Standard BioBrick...visit iGEM: BioBrick (RFC 10) Back to Top BioBrick BB-2 (RFC 12) Prefix: GAATTC GCGGCCGC T ACTAGT G Prefix...XbaI, SapI For more info, visit iGEM: BioBrick BB-2 (RFC 12) Back to Top BglBrick – Berkeley Standard ...assembly, even as more parts are added. Therefore, the same reaction which ligates a promoter into an empty ...

...sites on the same chromosome. Most multiplex systems enable researchers to target anywhere from 2–7 genetic...protein, instead of a two-component system (Figure 9C) SAM activators - co-expression of dCas9-VP64 with a modified...left free to interact with the target DNA. Figure 2: Overview of the NHEJ repair mechanism Cas9 will only...Cas9 is included in the gRNA-containing plasmid, or 2-vector systems, in which Cas9 must be delivered separately...your experimental cell population (Figure 8E). In a 2-vector system, you’ll need to either co-infect with...the presence of infectious organisms (like SARS-CoV-2 ) and genetic mutations. Similar to Cas9 and Cas12...Cas enzymes? CasPEDIA is an encyclopedia of Class 2 CRISPR systems with wiki entries describing enzyme...

...cells; function is unknown Monomer with a valency of 2 IgE Protects against parasites and is responsible ...such as anaphylactic shock Monomer with a valency of 2 IgG The most abundant isotype in blood, also found...secondary immune response Monomer with a valency of 2 IgG1; IgG2; IgG3; IgG4 IgM Produced during the primary... ranging from antibody fragments (shown in Figure 2) to completely unrelated molecules. Examples include...used for assembling protein nanostructures. Figure 2: An antibody (IgG, far left) compared to alternative...application Collect and lyse samples to make proteins available. Tissue samples may need additional processing...culture samples to extract proteins. Homogenize (tissues only) and lyse. Cross-link the samples with formaldehyde...

...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...targeting multiple loci can be easily expressed in the same cell or organism. The ability to precisely target...

...phage display. J Immunol Methods. 2000 Aug 28;242(1-2):159-81. Carlos Barbas Vector system for expression...+J phagemid libraries. Immunol Lett. 2015 Oct;167(2):95-102. Joanna Bereta Sybody Generation Toolbox for...microscope, or detect when the protein is present in a sample--just to name a few. Traditional sources of antibodies...

... Testing Samples Small samples of control AAV vectors for trial purposes in serotypes AAV1, 2, 5, 8, 9...Service overview. Function Chemogenetics DREADDs, PSAMs Optogenetics Channelrhodopsins, Halorhodopsins, ...

...referred to as intermolecular or bimolecular FRET) or (2) conformational changes within a protein where the...standard consisting of ECFP and EYFP connected by 2 GGSGGS repeats pET28CLY3 Peptide linker standard consisting...the donor and acceptor are both fused to the same protein (also referred to as intramolecular or unimolecular...

...Cre-ERT2 CAG Mammalian Heller 112622 pVHC_PGKneoLox2DTA.2 Venus, Cre-ERT2, targeting vector with MCS for homology...homology arms Mammalian Heller 112623 pEHC_PGKneoLox2DTA.2 Emerald, Cre-ERT2, targeting vector with MCS for homology... arms Mammalian Heller 112624 pmTHC_PGKneoLox2DTA.2 TFP, Cre-ERT2, targeting vector with MCS for homology...arms Mammalian Heller 112625 ptdTHC_PGKneoLox2DTA.2 tdTomato, Cre-ERT2, targeting vector with MCS for ...25;1504(4):467-86. doi:10.1016/0022-2836(81)90375-2. PubMed . Ventura A, Meissner A, Jaenisch R, Jacks...translocation. Inversion: If the loxP sites are on the same DNA strand and are in opposite orientations, recombination... is reversed. Deletion: If the sites face in the same direction, the sequence between the loxP sites is...

...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...