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We narrowed to 986 results for: IGH@;

Showing: 561 - 600 of 986 results

  2. Design Tips for Prime Editing

    Type
    Blog Post
    ...location bind to Cas9 with canonical sgRNA structure (right). Created with BioRender.com. What to edit The...3 or more mismatched bases can often evade MMR (right). Created with BioRender.com. For specific prime...prime editors bound to sgRNAs only drive nicking (right). Start with nicking sgRNAs ~50 bp from the prime...2024). Figure 9: epegRNAs (left) and PE7 (right) offer two different strategies for protecting pegRNAs...

  3. CRISPR 101: Targeting Non-Coding RNAs with CRISPR/Cas9

    Type
    Blog Post
    ...genome is made of protein-coding genes, and you might be wondering what the rest of our genome could possibly...non-coding RNAs! You probably didn’t see that coming, right? Non-coding RNAs (ncRNAs) are transcribed but do...versatile option. It can be used to target lncRNAs with high specificity, particularly in applications requiring...Hazan J, Bester AC. CRISPR-Based Approaches for the High-Throughput Characterization of Long Non-Coding RNAs...

  4. Plasmids 101: The Wide World of Natural Plasmids

    Type
    Blog Post
    ...just for lab specimens — there is evidence that lightning can also promote gene transfer in soil bacteria...microbial offense One way to triumph over your neighbors is to eliminate the competition, and Penicillium...specialized degradative capabilities are often tightly regulated to be active only in the presence of ...Simonet, P. (2001). Laboratory-scale evidence for lightning-mediated gene transfer in soil. Applied and Environmental...

  5. Making CRISPR Plasmids Using Fragmid

    Type
    Blog Post
    ...If you’ve ever been looking for just the right CRISPR vectors on Addgene and found instead ones that ...Doench reports even the greenest members of his lab, high school interns, were able to use Fragmid to design...making it easier to get the vectors that are just right for your experiment. As always, we recommend checking...components by clicking the blue text in the upper right-hand corner.      Step 3: Reviewing the ...

  6. Cpf1 Update: Comparison to Cas9 and NgAgo

    Type
    Blog Post
    ...NGG) in both distribution and GC-content, which might improve editing in AT-rich genomes. Additionally...editing efficiency for the Cpf1 orthologs was only slightly lower than that of the widely used SpCas9 and ...that AsCpf1’s off-target rate is similar to that of high fidelity Cas9s eSpCas9 and SpCas9-HF1. Both AsCpf1... cleaves relatively far away from the PAM, NHEJ might retain the PAM site. Therefore, if HDR does not ...

  7. Plasmids 101: Terminators and PolyA signals

    Type
    Blog Post
    ...elements. A high affinity terminator may be desired for multi-cistronic constructs where high termination...terminators, such as T7 and rrnB, as well as engineered high-efficiency terminators such as T0. Rho-independent...poly-uracil tract as shown in the figure to the right. The tertiary structure of the hairpin-DNA complex...

  8. Tips for Titering Your Lentiviral Preps

    Type
    Blog Post
    ...RNA. Measuring p24 levels in a viral prep is straightforward and rapid as there are several commercially...Studies have found that values can be 10-1000 fold higher than for functional assays depending on the vector... collected lentiviral prep may be significantly higher than the same prep that has been stored in the ...routinely produce virus may want to choose more straightforward approaches such as the FACS-based or colony-forming...

  9. Mobile-CRISPRi: Bringing CRISPRi to Diverse Bacteria

    Type
    Blog Post
    ...the Mobile CRISPRi cassette flanked by left and right Tn7 end sequences. Donor bacteria with a chromosomal...scheme. Mobile-CRISPRi integrates downstream of the highly conserved glutamine-fructose-6-phosphate aminotransferase...cell, the CRISPRi DNA (purple) flanked by left and right Tn7 end sequences (green) integrates downstream ...mirabilis, had only ~1 transfer per 1,000,000 colonies. Higher transfer rates are better suited for genome-wide...

  10. Starter guide to induced pluripotent stem cells (iPSCs) part 1:  A renaissance in regenerative medicine

    Type
    Blog Post
    ...of them: Characteristic ESC ASC Self-renewal High Low once isolated from the human body Usage Universal...specific cell types can be produced Ethical concern High. Embryos are destroyed during ESC isolation. Low...post-transplantation rejection when used therapeutically High Low (if ASCs are isolated from the patient) For...each other in many aspects. Like ESCs, iPSCs have a high self-renewal rate and can be used universally to...

  11. Fluorescent Proteins 101: Fluorescent Protein Timers

    Type
    Blog Post
    ...organelle, and protein localization. For instance, you might dye your cells and look at them under a microscope...population of timer indicates the age of the pool: the higher the ratio, the older the pool. FP timers have ... al. Protein Excitation (nm) Emission (nm) Brightness pKa Maturation Plasmids Fast-FT 403 (blue),...sections are formed. In synthetic biology, FP timers might be able to tell researchers whether old or new cells...

  12. Transferable Skills: Negotiation

    Type
    Blog Post
    ...At Addgene, we've set out to fill this gap by highlighting that researchers learn many transferable skills...the lab and by offering advice on areas where you might need some help. Today in our transferable skills...Although this approach to negotiation is the most straightforward, it can mean that everyone leaves disappointed...

  13. Degrading DNA with Cascade-Cas3

    Type
    Blog Post
    ...Cas effectors to light. One lesser-known Cas is waiting for its chance in the spotlight — Cas3. Cas3 can...Cas effectors can’t, while maintaining relatively high efficiency. These alterations are achieved through.... E., Bravo, J. P., Ramos, D., Hibshman, G. N., Wright, J. T., & Taylor, D. W. (2023). Structural snapshots...

  14. Plasmids 101: Degron Tags

    Type
    Blog Post
    ...added to media, buffer, etc., which will bind and light up a tagged protein for imaging. Given its popularity... like the SMASh tag does. However, it is a straightforward system – HaloTag and its PROTAC are all that...tag Are all tags created equal? No! Choosing the ‘right’ tag does depend a lot on your unique experimental...function is easily perturbed by tags, then a small tag might be the way to go (dTAG/SMASh). If you don’t want...

  15. Google Forums Round Up: First Impressions of NgAgo

    Type
    Blog Post
    ...but, to start, let's review why NgAgo is in the spotlight and take a moment to remember that NgAgo has only...a single experiment. Also, the NGG PAM sequence might not always be available at an optimal distance from...mean the technology is dead in the water. On the bright side, successful indel generation and epitope knockin...supposedly alien 5’-phosphorylated ssDNA guides, which might be rapidly degraded by the cellular machinery, leaving...

  16. Genome-wide Screening Using CRISPR

    Type
    Blog Post
    ...being used to perform genome-wide screens and highlight some of the reagents that have been made publicly... library. Choosing the CRISPR library that is right for you There are several factors to consider when... Findings from this type of experiment can shed light on the mechanism by which cells gain resistance ...

  17. Plasmids 101: Multicistronic Vectors

    Type
    Blog Post
    ...are studying a novel gene, is not always a straightforward process. Rather than try to directly detect...cells that are expressing your gene of interest at high levels. Unlike vectors that express screenable or.... (J Virol. 1988 Aug;62(8):2636-43.) PubMed. 3. Highly Efficient Multicistronic Lentiviral Vectorswith...):845-60. doi: 10.1089/hum.2008.188.) PubMed. 4. High cleavage efficiency of a 2A peptide derived from...

  18. Tips for arabidopsis transformation

    Type
    Blog Post
    ..., so it is pretty important to get this process right the first time. I’ll lay out some guidelines here...transformation process The process itself is straightforward. When your agrobacterial cultures are ready...s flowers inside. 50 mL falcon tubes or plastic weigh boats are both great options for this, depending... et al. (2007) ‘Improved Gateway Binary Vectors: High-Performance Vectors for Creation of Fusion Constructs...

  19. Great Results Start with Great Standard Curves

    Type
    Blog Post
    ...the unknown sample’s measurement is too high, start with a higher concentration of standard or make smaller...range is the linear span between the lowest and highest concentrations that the curve can accurately measure...labmates to get a sense of the concentrations you might expect to see for the unknowns in your experiment...

  20. CRISPR 101: Making a Knock-In Cell Line

    Type
    Blog Post
    ...cell cycle rates; rapidly diving cells will have higher HR frequencies than quiescent cells. It is important...ends of the ssDNA to find a repair template. The highest HDR efficiency is observed when inserts are within... need to incorporate homologous sequence to the right and/or left of the knock-in site. The length of ...: 387-390 (2020). 10.1016/j.jbiotec.2019.11.010 Wright, D. W., Shah, S. S., Heyer, W. D. Homologous recombination...

  21. Minigenomes - a Safe Way to Study Dangerous Viruses Like the Ebola Virus

    Type
    Blog Post
    ...contributed by guest blogger Tessa Cressey. The highly pathogenic Ebola virus belongs to the group of ...necessity for working with this virus under the highest biosafety level conditions, BSL-4. In this regard..., it is important to choose cell lines that are highly transfectable.  The amounts of each plasmid transfected...vary between labs, initial titration experiments might be helpful to achieve optimal results. It is important...

  22. Tips from the Repository Trenches: Using Barcodes to Track Samples

    Type
    Blog Post
    ...samples that come through our doors. It would be a nightmare if we didn’t have a simple way to track all of....2%) transform the first time and can be stored right away. Every week, the lab performs an average of...bacterial glycerol stocks on Fridays. From each overnight culture, we prepare three sets of 2D barcoded ...for a way to organize samples within your lab, we highly recommend using barcodes! Barcoded tubes are easily...

  23. Sonic Hedgehog? Sleeping Beauty? Learn About the Genes Behind Addgene’s Conference Room Names

    Type
    Blog Post
    ...where the brain fails to divide properly into the right and left hemisphere. Addgene has several plasmids...Spaghetti squash (Sqh) is the Drosophila Regulatory Light Chain (RMLC) of nonmuscle myosin II (Karess et al...PMC2395242. Jordan, Pascale, and Roger Karess. "Myosin light chain–activating phosphorylation sites are required...12490562. Karess, Roger E., et al. "The regulatory light chain of nonmuscle myosin is encoded by spaghetti-squash...

  24. Lab to Office Culture Shock

    Type
    Blog Post
    ...times during a 10 minute conversation. Leaving a tight-knit team like that can and will be a little gut-wrenching...camaraderie again in a non-academic career. You might not be in a lab anymore, but you will very likely...academia provide a formal training regime and the right set of tools to help you do your job as well as ...lab whether you realize it or not. These skills might include collaborating, managing, and communicating...

  25. How to Write a Scientific Review Article

    Type
    Blog Post
    ...that each paper taught me a few things and also highlighted a few dozen things that I didn’t know about. ... whatever time you have. If you’re like me, you might be surprised at how much you can get done between...article. This is normal and it means you’re on the right track!  Get feedback After you’re done editing, ...

  26. Mesothelioma - Causes, Symptoms, and Treatment

    Type
    Blog Post
    ...While there is a dose-response relationship (the higher the exposure, the greater the risk of disease) ...developments recently is a new blood test for HMGB1, the high mobility group box 1 protein. Normally localized... test to see how well treatments are working to fight the cancer.  Beyond early modifications to standard...mechanisms) that work through body’s immune system to fight cancer. However, there are many different forms ...

  27. New CRISPR Tools: Cas7-11 and PASTE

    Type
    Blog Post
    ...team originally suspected that this new Cas7-11 might be phylogenetically classified as the more complex...Site-specific Targeting Elements (PASTE). Finding the right integrase for this new approach proved a challenge...is more specific and efficient than HITI and has higher purity of insertion than both HITI and HDR. PASTE...indels and no off-target effects, giving PASTE a high-fidelity rate. Both Cas7-11 and PASTE open up multitudes...

  28. Viral Production at Addgene

    Type
    Blog Post
    ...optimized for each specific virus in order to generate high quality preparations. After production, all virus...obtained using ddPCR are generally two to three fold higher than those achieved using the standard qPCR method...performed by ddPCR.  Quality Control Addgene ensures high quality viral vectors by optimizing and standardizing...staining or SYPRO Ruby staining and the molecular weight and relative intensity of the viral capsid proteins...

  29. How to Be an Excellent Trainee

    Type
    Blog Post
    ...choosing a quality mentor, and rightfully so! They can provide you with high-level career advice, open doors...won’t be very proficient afterwards. Below we highlight how to prepare and things to keep an eye out for...

  30. Harnessing TnpB for Genome Editing: A Compact and Versatile Tool for Your Research

    Type
    Blog Post
    ...CRISPR-associated nucleases are Cas9 and Cas12a, the heavyweights in the genome editing toolbox. However, both...particularly via viral vectors. While viral delivery is a highly effective method for gene editing and allows targeting... targeting mechanism. Small RNA elements called right-end RNA (reRNA) or omega RNA (ωRNA) guide the TnpB...structural scaffold derived from the transposon's right-end element, and a 20-nucleotide (nt) guide sequence...

  31. Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist’s View

    Type
    Blog Post
    ...neuronal networks Double inverted vectors expressing light inducible ion channels (plasmid # 20298 for example...manipulations in vivo, FLEx vectors expressing the light activatable ion channel under the control of Cre...proteins (e.g. postsynaptic PSD95-mcherry) are used in high concentrations in combination with low concentrations...vectors expressing the required recombinase. The high concentration of FLEx vectors guarantees that most...

  32. Tips for CRISPR Gene Editing in Mice

    Type
    Blog Post
    ... intensity was classified into four groups (4; brighter than control, 3; same as control (Cetn1/sgRNA1...sgRNA1), 2; darker than control, 1; very dark). Brighter fluorescence indicates a more successful gRNA design...your cells under a fluorescence microscope. The higher the amount of fluorescence, the better that gRNA...been inserted or deleted. Each lab will use a slightly different method to genotype their pups. Taking...

  33. The Challenges of Cell Culture

    Type
    Blog Post
    ... viruses to study. Interestingly, due to certain high profile problems surrounding cell culture over the...but these days it’s never been easier to conduct high quality work in cell culture. The problems with ... Fortunately it has never been easier to conduct high quality cell culture. Biorepositories around the...AstraZeneca-MedImmune, and Infors & Applikon. Due to his high level of experience he now travels around Europe...

  34. Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

    Type
    Blog Post
    ...developed, which differ in the number, stability, and brightness of the fluorescent proteins used in the system...Brainbow 3.0 (Cai et al., 2013), incorporates the highly photostable fluorophores mOrange2, EGFP, and mKate2...system to boost XFP expression. The increased brightness of Tetbow allows this system to be used to trace...Sakaguchi, Richi, Marcus N. Leiwe, and Takeshi Imai. "Bright multicolor labeling of neuronal circuits with fluorescent...

  35. Plasmids 101: Biotinylation

    Type
    Blog Post
    ...Biotin protein ligase (BPL) is responsible for this highly specific post-translational protein modification...photoreactive arylazide group. Under strong visible light, the arylazide group becomes reactive and forms ...: 20353954. 5. Kay, B., Thai, S., & Volgina, V. “High-Throughput Biotinylation of Proteins.” Methods Mol...PMID: 2388593 7. Holden, M. A., & Cremer, P. S. “Light activated patterning of dye-labeled molecules on...

  36. Replacing Paper: Tips for Choosing an Electronic lab Notebook

    Type
    Blog Post
    ...news briefing, to quick notes, to finances, to flights & ordering food - everything is covered.   So why...for a large institution or company, site licenses might be available as well. In most cases this applies...are collaborating with other labs or clients, you might need share data with them including reports and ...does the ELN provider protect and respect your IP rights and data ownership? IP and data ownership should...

  37. Four Ways to Package Transgenes That Exceed the Size Limit of Adeno-associated Virus

    Type
    Blog Post
    ... issue of “tiny vector, big transgene” by just straight up packaging an oversized genome of ~9 kb into...Generally an overlapping split vector has equal or higher transgene expression when compared to using a single...of non-endogenous sequences, such as a 77bp long highly recombinogenic homology sequence derived from the...method While the hybrid dual vector system has a slight advantage over the overlapping and trans-splicing...

  38. Tips for Getting a Faculty Position

    Type
    Blog Post
    ... can find his original blog post below! --Ed.  Eight years ago, I decided to write a "how to" manual ... and mentoring to several people that found my insights helpful. All went on to get faculty positions ...potential colleagues study. Think about how you might collaborate or, at the absolute minimum, can appreciate...free!) contains strategies, hints, anecdotes, and insights. After I wrote the first version of my manual,...

  39. CRISPR 101: Validating Your Genome Edit

    Type
    Blog Post
    ...donor, you can deliver a silent mutation in a neighboring nucleotide along with your desired edit. This...provided you have some idea of where in the genome you might expect off target editing to occur. Luckily, in ...you to Sanger sequence a manageable number of the higher probability locations. If you’re worried about ...Bombieri N, Giugno R, Pinello L. CRISPRitz: rapid, high-throughput and variant-aware in silico off-target...

  40. CRISPR 101: Engineering the Plant Genome Using CRISPR/Cas9

    Type
    Blog Post
    ...overview of plant genome engineering using CRISPR, highlight the specific modifications to CRISPR machinery...Lab.  As with all plasmids in the repository, we highly recommend reading the associated publications or...10.1104/pp.15.00636 Ma X, Zhang X, Liu H, Li Z (2020) Highly efficient DNA-free plant genome editing using virally...
Showing: 561 - 600 of 986 results