We narrowed to 718 results for: des.1

...Morgens et al. contains three types of guides: 1) targeting guides (blue), which target all protein-coding..., the toxicity of guides with off-target sites was profiled. Guides with exact or 1-bp mismatch off-targets...) or short (17-18 bp) guides. When comparing the toxicity of guides that have 1-bp mismatch off-target... Figure 1: Summary of the advantages of the new approach to genome-wide CRISPR screen design.   Bassik... in both screens, with the growth screen having a 1% false discovery rate and identifying >88% of previously...characteristics that influence gRNA off-target activity: 1) mismatches closer to the PAM are less tolerated than...Knockout Library are available from Addgene! References 1. Morgens, David W., et al. "Genome-scale measurement...

...occurs at a much lower efficiency than NHEJ. Figure 1: Overview of NHEJ and HDR following cleavage by Cas9...or search the tables below for CRISPR plasmids designed to introduce a DSB. To learn more about NHEJ, ... CRISPR Guide Viral Preps CRISPR Protocols gRNA Design Tools CRISPR Blogs CasPEDIA Content last reviewed...

...gene activation CRISPR/Cas9 Plasmid Variants Figure 1: Schematic of the CRISPR Collection. Plasmids 159654...can be designed using Benchling or using similar design tools and ordered as standard desalted oligonucleotides...phosphorylated oligonucleotides containing the guide target sequence: Up to two guides can be expressed...that utilize Golden Gate cloning to insert the desired guide RNA sequence by digestion of the backbone... #63798) Plasmids from Collection ID Plasmid Description PI Publications Kaufman, M. L., Goodson, N. B...

...Biology, 2(9), 537–541. DOI: 10.1021/sb400021j Figure 1: Multiplexing CRISPR knockdown to modulate expression...CRISPR guides. The first one binds to sfgfp with either 0, 10, 11, 14, or 20 matching nucleotides, the ...also like... CRISPR Guide CRISPR Protocols gRNA Design Tools CRISPR Blog Posts The David Bikard Lab at...mcherry with either 0, 11, 18, or 20 matching nucleotides. Together, these plasmids allow to explore the...

...research and innovation programme and EFPIA. Figure 1: Representation of RESOLUTE mission. The combination...proteomic interactions represent the framework that provides biological context. Assays monitoring impedance...sub-cloning into a wide range of Gateway-compatible pDEST vectors. ID Plasmid Gene/Insert Industry PI Pooled...solute carrier family of proteins. Library ID PI Description Human SLC Activation Library 132561 Superti-Furga...

...activation-induced cytidine deaminase (AID). Figure 1: Overview of CRISPR base editing. Browse, sort, or...protein (Mu- GAM). While many base editors are designed to work in a very narrow window proximal to the... CRISPR Guide Viral Preps CRISPR Protocols gRNA Design Tools CRISPR Blogs CasPEDIA Content last reviewed...

...~50% Table 1: Nanoblades editing efficiencies of primary cells. Comparing Nanoblades to Existing CRISPR...components into 293T cells. Figure 1: The key components of Nanoblades. Image from Mangeot et al., 2019....loaded with 1-4 different gRNAs. Viral envelope proteins To alter the cell tropism of Nanoblades, different... ~50-70% for these four cell types (Table 1). Additionally, hiPSCs and mouse BM cells still behaved like...mouse-model generation. Key components of Nanoblades Nanoblades are a Cas9 RNP delivery vehicle that’s based...) are expressed to pseudotype Nanoblades. The team found that Nanoblades pseudotyped with a mix of VSV-G...using Nanoblades? Plasmids are available from Addgene. Genome editing primary cells with Nanoblades Primary...

...for brain circuit mapping and connectomics. Figure 1: Reconstruction of a sparse set of neurons across ...light microscopy. In brain tissue, these protein barcodes allow for efficient machine-learning segmentation...enhancers for morphological readout. How to Use PRISM Barcodes are delivered as pooled AAVs either directly to...bias in packaging representation. To read out the barcodes, each protein tag can be imaged with a corresponding...combinations of protein bits. B) Multiplex imaging of barcodes with expansion microscopy and reconstruction of...Sheridan et al. (2025). PRISM Plasmids ID Plasmid Description Tags Additional Resources PRISM was developed...., & Payne, A.C. (2025). Combinatorial protein barcodes enable self-correcting neuron tracing with nanoscale...

...spike protein (MERS, CoV-1, CoV-2), modification, or variant. ID Plasmid Description Mutations Industry PI...138152 pLentiEGFPdestablized - EFS-EGFPd2PEST-2A-MCS-Hygro Lentiviral vector expressing destabilized EGFP...neutralizing activity are listed below: ID Plasmid Description PI 44965 NL4-3 mCherry Luciferase Lentiviral ...

...vectors. PLoS ONE, 9(8), 1–10. https://doi.org/10.1371/journal.pone.0106472 de Backer, M. W. A., Brans,...promoters. Gene Therapy, 28(1–2), 56–74. https://doi.org/10.1038/s41434-020-0169-1 Oh, J. Y., Han, J. H., ...goals when designing and testing your AAV.  Figure 2: Comparison of AAV serotypes 1, 2, 5, 6, ...the selection process. Factors to think about are (1) your experimental conditions and measurements; (2...serotype, promoter, and titer.     Figure 1: Flow chart depicting a) steps for selecting serotypes...injection site than AAV2 (Watakabe et al., 2015) (Figure 1). You’ll need to consider if such properties could...Image modified from Aschauer et al., 2013, Figure 1. Used under Creative Commons Attribution License. ...

...and interrogate with conventional reagents. Figure 1: Axon guidance cues and receptors regulate the development...IGDCC3 (PUNC) Antibodies ID Recombinant Antibody Description Isotype Reactivity Recommended Applications PI...signaling during development. The IPI collection includes anti-glypicans that accurately detect glypican...scientists worldwide. ID Recombinant Antibody Description Isotype Reactivity Recommended Applications PI...neurological disorders. The initial collection includes antibodies to Neurexin and Neuroligin (Link opens...Neuroligin Antibodies ID Recombinant Antibody Description Isotype Reactivity Recommended Applications PI... Integrin Antibodies ID Recombinant Antibody Description Isotype Reactivity Recommended Applications Return...

...cells will still appear green despite having active Cas9 (see tip 2). Pro-Tips 1. To quantify Cas9 activity...gRNA-transduced samples (data not shown). Pro-Tips: 1.Designing primers against genomic DNA to amplify the gRNA...cancer cell line in about 1 month, which I’d say is pretty amazing. Despite the shifting landscape of ...Materials Used 1. lentiCas9-Blast (Plasmid #52962) was a gift from Feng Zhang and is described in Improved...The entire process from start to finish took about 1 month. Cas9 delivery Overview: Before any editing...lentiviral vector, lentiCas9-Blast (52962-LV, Figure 1). Some cells were then used to generate a monoclonal...rest of the cells were frozen down. Figure 1: Cas9 expression in A549 cells transduced with lentiCas9...

...1774 for neomycin selection. Bob Weinberg 10676 pMKO.1 GFP MoMLV U6-driven plasmid for shRNA expression with...contain a viral genome encoded on RNA. The genome includes a reverse transcriptase and an integrase to insert...plasmids that have inserts. ID Plasmid 5'LTR Description PI 1780 pBABE-neo largeTcDNA MoMLV Expression...Gamma-Retroviral Packaging Plasmids ID Plasmid Description PI 8449 pUMVC Contains MoMLV gag and pol . Use...Gamma-Retroviral Envelope Plasmids ID Plasmid Description PI 1733 pCI-VSVG VSV-G recommended for the FELIX...

...vectors. BMC Biotechnology, 6(1). https://doi.org/10.1186/1472-6750-6-1 O’Brien, J., Hayder, H., Zayed...endogenous miRNA machinery for processing (Figure 1). The shRNA sequence is transcribed to produce hairpin... recruits other silencing proteins. Figure 1: Comparison of shRNA and miRNA processing and silencing...,"rgb":"rgb(0, 82, 169)","rgba":"rgba(0, 82, 169, 1)"},"button_font":{"size":19,"styles":{"font-family...:"rgb(255, 255, 255)","rgba":"rgba(255, 255, 255, 1)"}}{% endraw %}{% end_module_attribute %}{% module_attribute...297. https://doi.org/10.1016/s0092-8674(04)00045-5 De Rie, D., Abugessaisa, I., Alam, T., Arner, E., Arner...Gebhard, C., Goldowitz, D., Guhl, S., Ha, T. J., . . . De Hoon, M. J. L. (2017). An integrated expression atlas...

... Caltech and can help guide your decision. Figure 1: Decision tree for choosing your AAV capsid. Your ...tdTomato Control Boyden 83900 pAAV-mDlx-GFP-Fishell-1 Dlx GFP Control Fishell 99130 pAAV-mDlx-NLS-mRuby2...at the bottom of this page. For more in depth descriptions of each serotype, check out Addgene's blog on...Browse Available PHP.eB AAV ID Name Promoter Description Category PI Controls 28306 pAAV-FLEX-tdTomato... Browse Available PHP.S AAV ID Name Promoter Description Category PI 28306 pAAV-FLEX-tdTomato CAG tdTomato...Browse Available PHP.V1 AAV ID Name Promoter Description Category PI 104052 pAAV-CAG-DIO-EYFP CAG EYFP...Browse Available MaCPNS1 AAV ID Name Promoter Description Category PI 37825 pAAV-CAG-GFP CAG GFP Control...

... nucleases. Genome Res. 2010 Jan;20(1):81-89. PubMed. 2. Bauer DE, Kamran SC, Lessard S, Xu J, Fujiwara...harness non-homologous end joining repair (NHEJ) [1]. Our initial work was with TALENs, in collaboration...from the University of Pennsylvania.     References: 1. Lee HJ, Kim E, Kim JS. Targeted chromosomal deletions...11;342(6155):253-257. PubMed. 3. Canver MC, Bauer DE, Dass A, Yien YY, Chung J, Masuda T, et al. Characterization...and Bauer lab's related paper describes their methods for CRISPR design, cloning, and delivery, as well...

...This gives each guide a score ranging between 0 and 1. The cutting frequency determination (CFD score) is...converted to a weight (which ranges between 0 and 1) so that they can be used in the rank equation above...converted to a weight (which ranges between 0 and 1) (shown in the graph below) so that it can be used...experiment in the program Design gRNAs by inputting the target regions Design donor RNA Design primers Review and...time for this project was spent selecting guides, designing primers, and running analyses. New team members...originals if desired. It works as a whole or in part. For example, you can analyze experiments designed outside...CrispyCrunch, or you can input pre-existing guides for primer design. The ecosystem of CRISPR tools Before...

...acceptable. Aims should be mechanistic, doable by 1-3 people in 1-5 years, and contribute to resolving the overarching...possible, do at least two full length practices (~1 h) and get feedback. It’s also helpful to practice...the knowledge gap? This includes the overview and the individual aims. Describe the question, the bare ...often hear, "I have an interview next week and it includes a chalk talk. What should I include in my chalk...may not actually help. Ultimately, if you cannot describe the likely outcome with some specifics, the search...cannot either. Therefore, you need to be able to describe experimental approaches in terms of the question...approach, anticipated outcomes, and preferably a description of the payoff if the aim is successful. Note ...

... your account. We will notify you by email within 1 business day regarding your account verification. ...Plasmid Gene/Insert PI Kits Kit name Type PI Description High Complexity Golden Gate Assembly Standards...PDB-deposited), disulfide-free, mesophilic, and designed to be diverse in topology, size, and isoelectric...Pooled Libraries Pooled Library name Type PI Description HR700_TP53 Exon Mutation Libraries CRISPR Thorsten...Parkinson's Disease; For target validation and drug design Antibodies Addgene distributes ready-to-use recombinant...materials and intellectual property rights, and provides liability protection for material providers. Addgene's...

...cell surface proteins. J Am Chem Soc. 127(37):12770-1. PMID:16159249...label of their choice according to the methods described by George et al. (2004). The ACP and SFP Synthases...respectively) may be expressed and purified as described previously in Stachelhaus et al. (1998) References...

...References Akhtar, W., De Jong, J., Pindyurin, A. V., Pagie, L., Meuleman, W., De Ridder, J., Berns, A....evaluate all of the pegRNAs in one experiment (Figure 1). Through this method, we found key determinants of...diverse edit types and cell contexts. Figure 1: High-throughput screening of pegRNA efficiency and...Article 3. https://doi.org/10.1038/s41576-022-00541-1 Kim, H. K., Kim, Y., Lee, S., Min, S., Bae, J. Y.,...DNA repair determinants. Nature Biotechnology 2023, 1–11. https://doi.org/10.1038/s41587-023-01678-y Li,...editing systems in multiple cell types. Cell, 186, 1–17. https://doi.org/10.1016/j.cell.2023.03.034 Resources...this means we could design hundreds to thousands of different pegRNAs for each desired edit, it simply becomes...

...formulation buffer is phosphate-buffered saline containing 1 mM sodium azide. This buffer is not compatible for...protocols, titering methods, and quality control are described below. Information for a specific antibody can...

...available as a substrate for another round. Figure 1: Overview of prime editing. Created with BioRender.com.... The longer pegRNA also encodes a primer binding site (PBS) and the desired edits on an RT template. ...post . Need help designing a pegRNA? Try the DeepPrime (Link opens in a new window) design tool from Hyongbum...pegRNA). As with a typical gRNA, the pegRNA is designed with a spacer that binds to a specific genomic...3’ flap binds to the PBS of the pegRNA and the desired edit is incorporated into the DNA by reverse transcription... CRISPR Guide Viral Preps CRISPR Protocols gRNA Design Tools CRISPR Blogs CasPEDIA Content last reviewed...

...thanks to our guest blogger, Harshana S De Silva Feelixge. Harshana S De Silva Feelixge is a researcher whose... post was contributed by guest blogger Harshana S De Silva Feelixge. Gene therapy technologies hold ...reconstitution with a normally functioning T cell repertoire (1). In this example, it was possible to transduce the...significant improvements in vision over a period of 1 to 3 years – the treatment appears to be both safe...research and science communication.     References 1. Gaspar, H. B., S. Howe, and A. J. Thrasher. "Gene...form, translating gene therapies from conceptual design to clinical trials involve identifying a therapeutic...

...HIV-1 were transduced with a lentiviral vector containing an antisense sequence against the HIV-1 envelope... the transfer plasmid encoded the desired transgene flanked by HIV-1 long terminal repeats (LTRs) that...Trono and collaborators turned to the lentivirus HIV-1, well-known to infect non-dividing cells. The first...plasmids. The packaging plasmid carried a mutated HIV-1 provirus that couldn’t package itself due to a few... targeting and tracing cells in the brain (Figure 1). 2nd generation lentiviral packaging system The graphic...infectivity, is often used as a substitute for HIV-1 Env, which can only infect CD4+ cells. 3rd generation...lentivirus in the comments section below! References 1. In vivo gene delivery and stable transduction of ...

...software such as SPAdes, Shovill, apc, and NOVOPlasty. The pipeline contains three de novo assemblers,...and recorded as a DNA strand is synthesized (Figure 1). Sanger sequencing requires four separate reactions...trace” file to determine the sequence. Figure 1: Overview of Sanger sequencing technology. Created...manually checks and assembles the raw data.  Addgene’s de novo assembly pipeline usually generates the correct...raw FASTQ reads are used for either guided or new de novo assembly. If a full anticipated sequence is ... doesn’t resolve the assembly issues, we can redo de novo assembly within Geneious.  While this may seem...steps that gave us our initial incorrect assembly, de novo assembly within Geneious gives us additional...

... Oligos Oligo 1: Add 'tcaa' to the 5′ end of your forward guide sequence (see Figure 1). Oligo 2: Add... Figure 1). Perform standard phosphorylation and annealing of the two oligos. Figure 1: Components... gene editing through de novo induction of meristems. Nature Biotechnology, 38(1), 84–89. https://doi....step-by-step guide for using a Dra2TnpB. Step 1: Design the 20-nt Guide Sequence Identify a TAM site ... promoter for ωRNA (reRNA) expression (see Figure 1). Both plasmids contain a hygromycin resistance gene...protocols (pp. 245–260). https://doi.org/10.1007/978-1-0716-4358-7_20  Karmakar, S., Panda, D., Panda, S...1166–1187. https://doi.org/10.1038/s41477-021-00991-1  Qiao, J., Zang, Y., Gao, Q., Liu, S., Zhang, X.,...

...allele. For a 1 kilobase gene, since potential target sites occur ~1 in every 8 nucleotides, restricting...Vazquez F, Parts L, Boehm JS, Golub TR, Hahn WC, Root DE, Garnett MJ, Tsherniak A, Iorio F (2019) Agreement...770–788 . https://doi.org/10.1038/s41576-018-0059-1 Veres A, Gosis BS, Ding Q, Collins R, Ragavendran ...j.stem.2014.04.020 Yang L, Guell M, Byrne S, Yang JL, De Los Angeles A, Mali P, Aach J, Kim-Kiselak C, Briggs... approaches, and provide updates on how best to design gRNAs. A more extended discussion of these subjects...important to remember that the same is true when designing gRNAs for using CRISPR technology – the “best”...and sequence are important considerations for designing your gRNAs. For indels, it's not so important ...

...using a metric that we describe in the Materials and Methods section of our paper (1). Lastly, it is possible...Green, Alexander A., et al. "Toehold switches: de-novo-designed regulators of gene expression." Cell 159.4...recombinase-based state machines in living cells” (1), into a more straight-forward, concise explanation...an RSM is to sequence the register. In our paper (1), we allowed individual bacterial cells to form colonies... not. How might one implement RSMs? In our study (1), we implement RSMs in E. coli with two plasmids (...inputs to endogenous gene expression (2). Our paper (1) explains how to arrange attB-attP pairs on a register...performance data from the indicated figures in our paper (1)) Input Plasmid Output Plasmid Fig. 3A pNR64 pNR160...

...coming closer to this DNA targeting ideal. Figure 1: (A) Representation of a zinc finger (ZF) nuclease...components of the array recognize individual nucleotides. For over ten years, zinc finger (ZF) arrays ...that came out in the spring of 2011, the groups describe a set of customized plasmids that can be used ... a TALEN kit through Addgene. This kit, first described in Nature Biotechnology in August, uses a serial...in Nature Biotechnology early in 2011 and they described their kit for assembling TAL effector arrays in...effector and TALEN kits here . References Efficient design and assembly of custom TALEN and other TAL effector-based...

...and no viral progeny are produced (Figure 1).  Figure 1: Comparison of the life cycle of a lentiviral...2004). HIV-1 gene expression: lessons from provirus and non-integrated DNA. Retrovirology, 1, 13. https... cannot integrate it into the host genome (Figure 1). IDLVs were first discovered by researchers investigating...you might choose an IDLV over a lentiviral vector: 1. No risk of insertional mutagenesis Lentiviral vectors...Brugman, M. H., Pike-Overzet, K., Chatters, S. J., de Ridder, D., Gilmour, K. C., Adams, S., Thornhill,..., H. E. (1996). Human immunodeficiency virus type 1 integrase mutants retain in vitro integrase activity...Berry, C., Ecker, J. R., & Bushman, F. (2002). HIV-1 integration in the human genome favors active genes...

...-Integrin alpha 5 beta 1 [IPI-ITGA5/ITGB1.2] Integrin alpha-5,Integrin beta-1 Human, Mouse IgG1 Rabbit...-Integrin alpha 5 beta 1 [IPI-ITGA5/ITGB1.4] Integrin alpha-5,Integrin beta-1 Human, Mouse IgG1 Rabbit...specific integrins in cellular processes.   Figure 1: Integrins can shape into three conformations: (A)... 26 recombinant antibodies (rAbs) either produced de novo using yeast display technology or produced from...immunoprecipitation and immunohistochemistry. Table 1: IPI antibodies  Antibody Target Proteins Reactivity...Human IgG1 Rabbit Anti-Integrin alpha M [IPI-LM2/1] Integrin alpha-M Human IgG1 Rabbit Anti-Integrin...Human IgG2a Mouse Anti-Integrin alpha M [IPI-LM2/1] Integrin alpha-M Human IgG2a Mouse Anti-Integrin...

...available serotypes. ID Plasmid Capsid Description PI 112862 pAAV2/1 AAV1 AAV packaging plasmid, expressing...required for AAV viral replication. This collection provides a curated selection of transfer, packaging, and... packaging eligibility, and more. ID Plasmid Description PI Return to top AAV Packaging Plasmids Packaging...repository for AAV vector production. ID Plasmid Description PI Return to top Other Resources Addgene's Viral...

...across scientific disciplines. Scientific Data, 8(1), 1–11. https://doi.org/10.1038/s41597-021-00981-0 Wright...excellent at infecting our target neurons (Figure 1). However, this one set of experiments didn’t provide...yet none of that data was published.    Fig. 1: Testing of three different AAV serotypes (AAV1, 8...392–400. https://doi.org/10.1038/s41434-018-0028-5 de Backer, M. W. A., Brans, M. A. D., Luijendijk, M....purification and formulation. Molecular Therapy, 12(1), 171–178. https://doi.org/10.1016/j.ymthe.2005.02.021...Stimulation. Frontiers in Human Neuroscience, 14(December), 1–12. https://doi.org/10.3389/fnhum.2020.578687 Additional...of negative data.  Negative AAV data is highly desirable  In a survey we conducted to improve our Addgene...

... compounds with various antimicrobial activities (1). She has developed a fast endophyte screening system...Rodrigo Arturo Rivera Solís, Universidad Autónoma de Yucatán, Mexico Dr. Rodrigo Arturo Rivera Solí...us coordinate and promote the awards. References 1. Martinez-Klimova, Elena et al. “Endophytes as sources...antibiotic” Biochemical Pharmacology 2017 Jun 15;134:1-17. PubMed PMID: 27984002. 2. Bilan, Dmitry S. et ...renewable biotechnology products and biosensors for herbicides as well as developing yeast as source for dietary...technology to engineer plants that are able to destroy DNA viruses, similar to the anti-phage CRISPR effect...cloning kit) she intends to make yeast capable of desaturating fatty acids. Identification of Insecticidal ...

...plasma HIV-1, improving life expectancy and quality for HIV-1 patients. However, ART is not an HIV-1 cure. ...activated HIV-1, measured via HIV-1 promoter-driven GFP expression. In cell lines that can produce HIV-1 toxic...with HIV-1. For two HIV-1 strains, Cas9/gRNA lentiviral expression significantly reduced the HIV-1 copy number...HIV-1 transcription and destroy infected cells, the other using wild-type Cas9 to remove the HIV-1 genome...currently infected with the lentivirus HIV-1. Today, HIV-1 can be controlled with antiviral therapies ...take on HIV-1 in vitro, and what obstacles must be overcome for clinical success. ART and HIV-1 reservoirs...Cutting HIV-1 out of the genome with CRISPR/Cas9 Before the introduction of CRISPR, HIV-1 genome editing...

...postdoctoral researcher at Instituto de Investigación Biomédica de Salamanca (IBSAL) and one of the coordinators...prompted us to create Crowdfight.   Figure 1: Crowdfight is a community of scientists who believe...collaborator for a specific task, Crowdfight will: 1) Find an expert with the right skill; 2) Confirm the... illustrates the power of collaboration, which despite the competitiveness of cutting-edge research, is...physics. Even non-scientists, such as lawyers, designers and other professionals, joined the platform; ...

...regulation of endogenous RNA. J Biol Eng. 2014 Mar 1;8(1):7. doi: 10.1186/1754-1611-8-7. PubMed.  Related Resources...targeting specific sequences of RNA is highly desirable for extending the complexity of genetic circuits...Zhao and colleagues, we now have just the tool…designer PUF proteins! A newly available PUF Assembly Kit...In a single step, researchers can now assemble designer PUF domains for RNA specificity engineering. “...proteins The tool she and her colleagues led by Zhao described and demonstrated in a report published in The ...translational repressor. In principle, however, designer PUF proteins can be fused to other components ...standard BsaI in place of the high fidelity version (despite what the original paper says). In their hands, ...

...for the wild-type. However, variants such as the P.1 (Gamma variant) and the B.1.351 (Beta variant) were... tested at the time of this study. Figure 1: Neutralization levels against spike pseudovirus decreases...lowest levels of cross-neutralization occurs in the P.1 and B.1.351 variants tested. Image from Garcia-Beltran..., Gosnell BI, Hanekom W, Moosa M-YS, Lessells RJ, de Oliveira T, Sigal A (2021) Escape of SARS-CoV-2 501Y.V2...

...unusual cases was either 1) a physiological effect of SARS-CoV-2 toward destabilizing one’s natural immune...an AdV transfer plasmid with the E1 and E3 (Early 1 and Early 3) genes deleted. These genes’ functions...certain contexts (Bauzon et al., 2003).        Figure 1: The adenovirus life cycle. Image reused from Kremer... review of the evidence. Emerg Microbes Infect, 8(1), 1679–1687. https://doi.org/10.1080/22221751.2019.1690953...adenoviral vector infected cells does not require de novo viral gene expression: Implications for gene...574–580. https://doi.org/10.1038/s41586-023-05949-1. PMID: 36996871. Tatsis, N., Fitzgerald, J. C., Reyes-Sandoval...viruses infecting humans (from seven species, designated as letters A–G) exist. The life cycle of AdVs...

...to determine purity of the AAV preparation. Figure 1: Silver staining of purified and non-purified AAV ...protocols, titering methods, and quality control are described below. AAV Production AAV distributed by Addgene...

... help!  General Design Figure 1: Two proteins are fused to the FP(11) and FP(1–10) fragments. When... Base FP Description Article (Link to Plasmids) PI Blue EBFP2 EBFP2(1–10) and Capri(1–10) for ...and 11 to yield large (1–10) and small (11) fragments. In several cases an FP(1–10) fragment may be compatible...Commun. 2017 Bo Huang mNeonGreen3 mNG3K(1–10) and mNG3A(1–10) for use with mNG2(11) Improved yellow-...sfCherry2 sfCherry2(1–10) and sfCherry2(11), also photo-activatable variant PAsfCherry2(1–10) Improved split...brings the pieces of the FP back together (Figure 1). This method is generally known as Bimolecular Fluorescence...even from a different base FP (for example, EBFP2(1–10) can be used with GFP(11)). Some of the articles...

...different plasmids to produce AAV: 1) the RepCap plasmid which provides the AAV replication (rep) and capsid...plasmid. This whole process takes ~3 hours to complete: 1 hours hands-on time and 2 hours for the qPCR run and... Special thanks for Dr. Dan Stone and Harshana de Silva Feelixge at Fred Hutchinson Cancer Center for...about AAV production and purification! References 1. Aurnhammer, C., Haase, M., Muether, N., Hausl, M.... repeat sequences. Human gene therapy methods, 23 1, 18-28. PubMed PMID: 22428977. 2. Choi, V.W., Asokan...replication; and 3) the transfer plasmid which encodes a transgene of interest that’s packaged into the...

...replication, OriP and the EBNA-1 sequence that encodes for the DNA binding protein EBNA-1 [7]. These two sequences...are based on the Epstein-Barr Nuclear Antigen-1 (oriP-EBNA-1) system where each vector contains a viral ...extra plasmid, pCXWB-EBNA-1 is used in the reprogramming cocktail. pCXWB-EBNA-1 lacks oriP and is unable...expressing additional EBNA-1 transiently. The transiently expressed EBNA-1 increases protein expression...pCXLE vectors listed below in Table 1 are available at Addgene. Table 1: List of pCXLE episomal plasmids,...obtained and the subsequent reprogramming success [1]. Fibroblasts need to be cultured and expanded in ...reprogramming factor cocktail, and viral protein EBNA-1. Methods for reprogramming hiPSCs from PBMCs While...

...type of choice in vitro is known as reprogramming [1]. The process can be divided into two stages: Dedifferentiation...//www.linkedin.com/in/kmukherjeephd/.  References 1. Hochedlinger, K. and R. Jaenisch, Nuclear reprogramming...from human peripheral blood. Cell Stem Cell, 2010. 7(1): p. 15-9. PubMed PMID: 20621044. PubMed Central PMCID...induced pluripotent stem cells. Hepatology, 2010. 51(1): p. 297-305. PubMed PMID: 19998274. PubMed Central... pluripotent stem cells. Nat Biotechnol, 2014. 32(1): p. 84-91. PubMed PMID: 24291815. PubMed Central ...keratinocyte lineage. Methods Mol Biol, 2014. 1195: p. 1-12. PubMed PMID: 24510784. PubMed Central PMCID: PMC4096605...induced pluripotent stem cells. PLoS One, 2010. 5(1): p. e8763. PubMed PMID: 20098701. 31. Tang, Z.H.,...

...Table 1 below outlines a few of the more common genetic changes found in E. coli strains. Table 1: Common...amber (UAG) stop codon by tyrosine insertion λ-thi-1 or thi1 Mutation in thiamine metabolism Requires exogenous... all based on E. coli K-12 and are considered BSL-1. Table 2: Lab strains of E. coli Strain Natural...plasmids, blue/white screening. F- endA1 glnV44 thi-1 recA1 relA1 gyrA96 deoR nupG Φ80dlacZΔM15 Δ(lacZYA-argF...) recA13 leuB6 ara-14 proA2 lacY1 galK2 xyl-5 mtl-1 rpsL20(SmR) glnV44 λ- JM109   General cloning and...strain for cloning repetitive DNA. endA1 glnV44 thi-1 relA1 gyrA96 recA1 mcrB+ Δ(lac-proAB) e14- [F' traD36...of an E. coli K-12 strain. F- λ- ilvG- rfb-50 rph-1 NEB Stable   For cloning into and storage of lentiviral...

...    Figure 1: A 1 mg/mL control is diluted in a series ranging from 1:2 to 1:32. Each individual...example, Figure 1 depicts a 2-fold dilution series. Each step of the series dilutes by 1:2 for a series...series that ranges from 1:2–1:32. The specific dilution series used will depend on the expected concentration... points fit the trendline. R2 ranges from 0 to 1 with 1 being a perfect fit and 0 indicating that there...individual step of the series is consistent at 1:2. When preparing a dilution series, use a new pipette tip for... the data points generated by replicate values provides useful information about the accuracy of the curve...If the unknown sample’s measurement is too low, redesign the standard curve dilution series with lower ...

...protein design goals, both from the Baker Lab and others. The introduction of de novo designed proteins...protein structures, called Top7. Figure 1: Top7 receives the Nobel Prize. Created with BioRender.com...concept that this kind of protein design is possible. Although Top7 was designed simply to be stable and novel...Hassabis and John Jumper! The Baker Lab focuses on designing entirely new proteins, improving our understanding...team was the first to develop and publish a human-designed protein that was not based on any existing protein...least that we’ve seen so far!). They started by designing a novel topology, then iteratively generated 3D...got closer and closer to achieving it. Once they designed the protein computationally, the researchers expressed...

...three-step approach for designing an experiment to identify the critical NANOG binding sites: 1. Find Enhancers...in combination with the techniques from Table 1, provides us with powerful GPS-like techniques to explore... chromosome, start, and stop. For instance (chr3, 1, 10) indicates a stretch of DNA at the very beginning...genetic elements like protein, RNA, and DNA (Table 1). As with GPS in the real world, coordinates alone...genomic coordinates into meaningful experiments. Table 1: An emerging family of "GPS for the genome" techniques...enhancers with the help of genome math. Figure 1: Genome math illustrated. Intersect (left) and set...histones with “GPS for proteins,” ChIP-seq from Table 1. Because of DNA’s 3D geometry and the chemical properties...

...recombineering system has three components (Figure 1): 1) Exo, 2) Beta, and 3) Gam. All three are required...frequency between 0.1% to 1%, and can be increased to as high as 25-50% by designing oligos that avoid activating...activating MMR: 1) use a strain of bacteria that has key MMR proteins knocked out or 2) specially design ssDNA ...  These changes can be 5’ or 3’ of the desired change.   Table 1: Overview of ds- and ss-DNA Uses in Recombineering...Considerations dsDNA Insertion or deletion of >20 nucleotides 1 out of 104 to 105 Use a high fidelity Taq polymerase... ssDNA Point mutations or changing a few nucleotides 0.1-1% or 25-50% when activation of MMR is avoided...from the 5’ end and generate 2 possible products: 1) a partially dsDNA duplex with single-stranded 3’ ...