...which had a total of 31 amino acid replacements relative to CFP484. Because the excitation and emission...combination results in a 2.6-fold increase in signal relative to mCerulean imaged with a standard CFP set. mTFP1...constructed 22 different mTFP1 fusions to both the C- and N- termini of the fluorescent protein and found... PubMed Central PMCID: PMC2292683. Shaner, Nathan C., et al. "A bright monomeric green fluorescent protein.... mTFP1 is also insensitive to physiologically relevant changes in pH. Find mTFP1 plasmids here! mWasabi...

...terminal); 2xNLS (C terminal) 47327 pET-28b-Cas9-His 6xHis (C terminal); NLS (C terminal) 62933...6xHis 6xHis (C terminal); NLS (C terminal) 62934 pET-NLS-Cas9-6xHis 6xHis (C terminal); NLS ...mouse. It’s also made targeted insertions relatively simple in C. elegans and bacteria. But CRISPRing typical...the stop codon!  Figure 3: How to design a C-terminal insertion with Easi-CRISPR. Potential guide...

...Michael, S., Kuleshova, E., Ojala, D., Huang, C.-C., Gerfen, C. R., Schiller, J., Dudman, J. T., Hantman,...our in-stock viral prep service.  AAV9 AAV9 has relatively broad tissue tropism and high transduction efficiency...doi.org/10.1089/hum.2008.123 Dalkara, D., Byrne, L. C., Klimczak, R. R., Visel, M., Yin, L., Merigan, W....scitranslmed.3005708 Goertsen, D., Goeden, N., Flytzanis, N. C., & Gradinaru, V. (2022). Targeting the lung epithelium...Ishikawa, Y., Jae, L. T., Wosen, J. E., Nagamine, C. M., Chapman, M. S., & Carette, J. E. (2016). An essential... S., Widrick, J. J., Messemer, K. A., Troiano, E. C., Moghadaszadeh, B., Peacker, B. L., Leacock, K. A...Horwitz, N., Beggs, A. H., Wagers, A. J., & Sabeti, P. C. (2021). Directed evolution of a family of AAV capsid...

...transversions C → TG → A A → GT → C C → G                   C → AG → C                   G → T A → C                   ...Rosser, S. J., Bi, C., & Zhang, X. (2021). Glycosylase base editors enable C-to-A and C-to-G base changes...deaminase could also perform cytidine (C) deamination, converting the C base to uracil (U). This is the same...given the pithier name CGBE1, for “C → G Base Editor 1,” and had a C → G editing efficiency of up to 68%...Beyond UNG: Optimizing C → G editors Now that several labs had shown that preferential C → G editing was possible...pyrimidines (C ↔ T).   Base transversion  Base changes from purine (A or G) to pyrimidine (C or T) or vice-versa...type of base editor invented. CGBE C → G Base Editor Converts C → G; also called a glycosylase base ...

...histidine amino acids on the N- or C- terminus of a protein. Neither the N or C terminus is preferable over ...binding of untagged proteins. Despite being a relatively low frequency amino acid, proteins containing...histidine residues expressed in frame on the N- or C-terminus of a recombinant protein of interest. The...need to decide whether to add the tag to the N- or C-terminus and how many histidine residues to include...or function of a single protein? Oftentimes, they rely on protein purification. In this strategy a single...contrast, the polyhistidine tag is very small and rarely affects protein function (Bornhorst 2000).  One...other affinity purification methods where binding relies on a specific protein conformation, the polyhistidine-metal...

...temperature of the PCR from 60° C to 61° C improved assay reliability. Different primers can have different...the ratio of genome-containing viral particles relative to the total number of viral capsids (which can...vectors consist of full particles (white arrowhead) relative to empty particles (black arrowhead). Scale bar...absolute titer values may not be as important as the relative titer values. Finally, because physical titer ... PubMed Central PMCID: PMC2957240. 5.Taylor, Sean C., Genevieve Laperriere, and Hugo Germain. "Droplet...that vary up to two-fold. How can we get more reliable AAV titering? For our viral service, we currently...vector preparations by qPCR. To ensure accurate and reliable results, we have optimized our assay in a few ...

.../000504294 Wang, T., Guan, C., Guo, J., Liu, B., Wu, Y., Xie, Z., Zhang, C., & Xing, X.-H. (2018). Pooled...rat, is another popular model organism. A close relative to the mouse, both are great mammalian models ...that make them an ideal model organism; they are relatively simple to maintain and breed and they have large...doi.org/10.13918/j.issn.2095-8137.2016.4.237 McCusker, C., & Gardiner, D. M. (2011). The Axolotl Model for ...elife.50651 Vieira, W. A., Wells, K. M., & McCusker, C. D. (2019). Advancements to the Axolotl Model for ...BSL-1 status, E. coli can be used as a model to relate these processes in more pathogenic bacteria (Blount...regeneration.  In addition, axolotls have many age-related phenotypic changes that are similar to those observed...

...for use in C. elegans. In 2017, the Korswagen lab developed Cas12a (Cpf1) for editing in C. elegans (Ebbing...editing from more sites of the C. elegans genome. Toolkits for editing in C. elegans The development of ...SapTrap-SEC kit for C. elegans genome engineering have also facilitated CRISPR editing in C. elegans. These...2018). To see more CRISPR tools for C. elegans, visit Addgene’s C. elegans CRISPR plasmids page. Future...Ruvkun for their seminal work on micro RNAs in C. elegans. The C. elegans work on small RNAs informed and drove... recovery of these edits. Initial work in C. elegans relied on selective markers such as drug resistance...possible in a wide range of organisms and cell lines. In C. elegans, one of the model organisms which I use to...

...bound to the C-terminal peptide. One of these C-terminal antibodies was Myc1-9E10 (aka anti-c-Myc [9E10]...sequences within the middle of the human c-Myc protein or the C-terminus, Evan, et al. isolated six different...that Myc1-9E10 and the other C-terminal antibodies only recognized the human c-Myc protein and did not recognize...experiments? Why Myc? When did the commonly used anti-c-Myc [9E10] antibody come into play? We’ll dive into...This protein had a highly specific antibody to the C-terminal five amino acids, meaning the tag could be...However, substance P has an amide group added to its C-terminus post-translationally, which turned out to.... As such, the peptide could only be added to the C-terminus of proteins of interest and proteins tagged...

...make your own. In my next post, I’ll explain the relatively straightforward process of generating your own...21356876. 5. J. Chow, Z. Marka, I. Bartos, S. Marka, J. C. Kagan, Environmental Stress Causes Lethal Neuro-Trauma...PubMed PMID: 28704652. 6. V. Rodríguez, D. Didiano, C. Desplan, Power tools for gene expression and clonal...Neurosci. 9, 703–709 (2006). PubMed PMID: 16582903. 8. C. J. Potter, B. Tasic, E. V Russler, L. Liang, L. Luo...

...37-16 °C cycling 4-base Bsal-HFv2 37-16 °C cycling 4-base BsmBI-v2 42-16 °C cycling... T4 DNA ligase 25 °C 1 hour 3-base or 4-base T4 DNA ligase 25 °C 18 hours 3-base or ... T4 DNA ligase 37 °C 1 hour 3-base or 4-base T4 DNA ligase 37 °C 18 hours 3-base or ...NEBridge® Ligase MM 37-16 °C cycling 4-base Bsal-HFv2® 37 °C Static 4-base Bsal-HFv2...Ligase MM 42-16 °C cycling 4-base BspQI, 1x NEBridge Ligase MM 42-16 °C cycling 3-base...Ligase Buffer 37-16 °C cycling 4-base PaqCl, 1x NEBridge Ligase MM 37-16 °C cycling 4-base...              37-16 °C cycling 3-base Sapl, 1x NEBridge Ligase MM 37-16 °C cycling 3-base ...

... 22nd International C. elegans meeting, I was again reminded of the extent that C. elegans researchers...and toolkits available at Addgene.  C. elegans open resources The C. elegans Neuronal Gene Expression Map...database specifically for C. elegans. This database provides detailed information about C. elegans 3’UTRs structures...The C. elegans community has always emphasized the need for open science and collaboration. The field... important for post-transcriptional regulation in C. elegans. To study the role of 3’UTRs, the Arizona...view the anatomy of worm layer by layer, the NeuroML C. elegans connectome, a model of the C.elegans neural...be used to simulate muscle tissue as it applies to C. elegans locomotion. To see the other resources included...

...genome engineering methods for C. elegans were released, signaling a new era in C. elegans research. Homology...of custom genomic modifications, is very robust in C. elegans, and the methods for HDR-mediated modification...the University of North Carolina has made CRISPR in C. elegans even easier - now, one can generate a fluorescent...-based gene tagging approach Multiple features of C. elegans have made CRISPR/Cas9 so successful in this...previously used a transgene selection strategy for their C. elegans/CRISPR experiments, but this strategy required... make it possible fluorescently label proteins in C. elegans, they inserted the SEC into a synthetic intron...system is both simple to use and highly efficient for C. elegans genome modification. Figure 2: Schematic...

...recognized by reliable antibodies. It is derived from the C-terminal region of the human proto-oncogene c-Myc. ... on the Anti-c-Myc [9E10] page for more details. Image credit: Addgene.   Find anti-c-Myc antibody...potassium channelrhodopsins Myc-tag antibody anti-c-Myc [9E10] now available! Improved voltage indicator...and tdTomato expressed layer 2/3 neurons in mouse. C) Action potential frequencies with (magenta) and without...10.1101/2021.09.17.460684.)   Myc-tag antibody anti-c-Myc [9E10] now available! by: Ashley Waldron The majority.... Since the generation of the first anti-c-Myc [9E10] (Even, et al., 1985), the Myc tag has become one...pulled up over 13,000 plasmids! Our recombinant anti-c-Myc [9E10] (isotype IgG2a) antibody is currently recommended...

... L L T C G D V E E N P G P P2A: (GSG) A T N F S L L K Q A G D V E E N P G P E2A: (GSG) Q C T N Y A...plasmid is packaged for viral delivery, or the relative expression levels between two or more genes is...ribosome skip the synthesis of a peptide bond at the C-terminus of a 2A element, leading to separation between...between the Glycine and Proline residues found on the C-terminus meaning the upstream cistron will have a ...the same mRNA. The term "self-cleaving" is not entirely accurate, as these peptides are thought to function... plasmids listed in the table above (and their related plasmids) are designed to have one or more of the...fluorescent reporter for easy detection.5  Just released: Addgene's Plasmids 101 eBook And now for some...

...PMC3223083. 6. McInnes, J. L., Forster, A. C., Skingle, D. C., & Symons, R. H. “Preparation and uses of...PMC164612. 11. Liou, Y.-R., Wang, Y.-H., Lee, C.-Y., & Li, P.-C. “Buoyancy-Activated Cell Sorting Using Targeted...Uses of biotin in molecular biology Biotin is a relatively small, water soluble molecule that does not interfere...fusion in which AviTag is added to either the N or C terminus of the protein. BirA has been shown to effectively...of some other common applications and links to relevant papers and webpages are listed below: EMSA (Electrophoretic...

... and C-C’) that give the feature its characteristic T-shape. The arrangement of the B-B’ and C-C’ palindromic... Tian, W., Liu, C., Lian, Z., Dong, X., & Wu, X. (2017). Deletion of the B-B’ and C-C’ regions of inverted...contains three palindromic sequences (A-A’, B-B’, and C-C’ regions) and forms a T-shaped structure. The Rep... Conatser, L. M., Lue, V. M., Dobbins, A. L., Li, C., Hirsch, M. L., & Samulski, R. J. (2020). Adeno-Associated... D. V. (2023). AAV- based vector improvements unrelated to capsid protein modification. Front Med, 10,...

...in fact, permeable to CO2 at low temperatures (-80°C) and once in the tube, the CO2 can alter the pH of...pH indicator phenol red. We froze samples in a -80°C freezer and then moved them to a closed dry ice storage...samples to controls that were only stored in a -80°C freezer and not exposed to CO2/dry ice. All samples...then tested whether returning the tubes to the -80°C freezer after storage on dry ice allowed the CO2 to... (Fig.4). To test this, we froze the tubes at -80°C and then moved them to closed storage on dry ice for...for 48 hours. We then moved the tubes back to a -80°C freezer for various recovery periods between 0 and... samples to controls that were only stored at -80°C. We then thawed the tubes after the indicated incubation...

...?" Many thanks to our guest, Darrell Kotton. Dr. Kotton is the David C. Seldin Professor in the Department...about Darrell, his research, and his focus on open science. Listen to the interview with Darrell Kotton...Introduction to Darrell Kotton and the Boston University Center for Regenerative Medicine. Darrell describes...low-risk but reliable projects. Chapter 9: Using stem cells as in-vitro models of disease. How Darrell is using...Darrell Kotton is the director of the Center for Regenerative Medicine at Boston University Medical School... School. Darrell’s research focuses on the lungs, and, among other projects, using stem cells to develop...develop in vitro models for lung disease. Darrell strives to promote open source biology in his own lab, ...

...editing toolkit to allow C to U edits using their RESCUE (RNA Editing for Specific C-to-U Exchange) system...RESCUE-S) offered ~76% on-target C-to-U editing while reducing off-target C-to-U edits by ~45% and off-target...lab published a paper on an RNA editor that allows C to U edits (RESCUE). We’ll walk through how these ...adjusted the structure of the guide RNA by introducing a C opposite the target A to be edited. This change specifies...residues that contact the RNA substrate resulted in 15% C-to-U editing activity. They then performed sixteen...cytidine deaminase activity. While RESCUE can perform C-to-U edits, the authors note that it retains its adenine... that RESCUE is most active in a 30 nt guide when C or U base-flips are present across from the target...