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...their experimental needs, and some of the common applications in the lab. Why Green Fluorescent Protein?...Re-engineering GFP to increase its range of color and application Almost as soon as its sequence was elucidated...the mutation definitions found in Table 2 may not apply. Table 1: The specific mutations comprising common...others) Neutral mutations    A multitude of applications Due to its size and ease of use, GFP and other...change. Typically, two fluorescent proteins with overlapping excitation/emission spectra are used; one fused... just scratched the surface of the potential applications for GFP. It has also been used to identify particular...

...these proteins are quite large — comprising approximately 1000–1400 amino acids — which poses a challenge...-strand breaks like those created by Cas12a. Applications of TnpB One of the major challenges in genome...), which is especially valuable for regulatory approval and public acceptance. Recently, viral vectors... is where TnpB offers a clear advantage. At approximately one-third the size of Cas9 or Cas12a, TnpB is...viral vectors. Even for advanced genome editing approaches like base editing and prime editing (Molla et...viral vectors and for multiplexed gene editing applications. As research into TnpB continues to evolve, ...

...This distinct approach means researchers can now select the best tool for their application from among many...incorporate A bases across from abasic sites. This approach resulted in a maximum A → T product purity of ...AYBEv3!). Although AYBE, AXBE, and ACBE do not approach the high product purity of CBEs and ABEs, they...). Several different teams succeeded with this approach around the same time (He et al., 2024; Tong et...on in the base editing field! From CRISPR-free approaches to mitochondrial base editing to base editors...

...performs well in many different antibody-based applications.  IPI’s recombinant version is a fusion of mouse...ligase cereblon (CRBN), including popular new approaches like PROTAC targeted protein degraders. But CRBN.... elegans embryo, where developmental processes happen within a few hours. In order to use the latest ...cellular processes in C. elegans: mScarlet3 for applications needing intense brightness (such as low-expression...simplicity and efficiency of this dual-plasmid approach makes it a valuable resource for advancing our...when the module unfolds under tension. When this happens, BiT and the LgBiT luciferase fragment (expressed...

...difficult to source a commercially available and appropriately conjugated antibody. While labs can conjugate...find,  it is often easier for labs to source appropriate antibodies for an indirect ELISA than for a direct... capture antibody must bind to distinct, non-overlapping epitopes so that they do not interfere with each...specific and can be difficult to develop if an appropriate conjugated antibody is not readily available....cross-reactivityChallenging to develop assay Competitive Appropriate for small moleculesReduced chance of cross-reactivity...choose the best ELISA for your needs. Good luck and happy binding! More resources on the Addgene blog Antibodies...

...hour. Find the SHERLOCK plasmids at Addgene! Applications of the SHERLOCK detection system The Zhang lab...portability. This method can be used for an array of applications including clinical diagnostics (e.g. pathogen...While the lab points out that the protocol is not approved for clinical use at this point, they hope that...patient samples. Find the DETECTR plasmids here! Applications of DETECTR in diagnostics DETECTR allows for...an array of molecular and clinical diagnostic applications. The company Mammoth Biosciences has been started...SHERLOCK detection system, DETECTR has not been approved for diagnostics yet, but is undergoing validation...

...Depending on the research question and downstream application, one or several of those techniques can be used...your flow analysis. However, certain downstream applications, like an in vivo adoptive transfer of those ...or free amines of proteins. Depending on the application, they can be membrane-permeant or membrane-impermeant...are positive or negative for the dye. The main applications of dye stains in flow cytometry are cell proliferation...with a high resolution for specific downstream applications, while maintaining a clean cell surface, dump...and bind DNA or free amines, depending on the application. Paul Heisig is a Research Associate in the lab...

...cells retain the same color, allowing for the fate mapping of cell populations over time. The ability to track...variety of species, cell types, and research applications. Labeling neurons with Brainbow The Lichtman...neurons in the cerebellum of mice, the team appropriately named their technology Brainbow. Brainbow 1.0...level of these XFP is not strong enough for all applications. To resolve this, similar systems have been ...cell type when crossed with mice expressing the appropriate Cre allele. The Confetti mouse contains the Brainbow...PMC3713494. Ghigo, Clément, et al. "Multicolor fate mapping of Langerhans cell homeostasis." Journal of Experimental...

...AAV11 ✓ ✓ AAV1 AAV1 was the first viral vector approved for gene therapy (Issa et al., 2023). It may be... 2016). This functionality makes it useful for mapping neural connections. Although derived from AAV2,...vessel barriers. It is especially popular for applications targeting liver tissue. In addition, it has ...the blood-brain barrier, making it useful for applications targeting the CNS (Issa et al., 2023). Additionally...treatment, which can pose problems for gene therapy applications. Cross-reactivity of AAV serotypes varies by.... A. (2023). Various AAV Serotypes and Their Applications in Gene Therapy: An Overview. Cells, 12(5), ...

...secondary antibodies.   Application-specific controls Some antibody-based applications have very specific ...then moving on to controls for antibody-related applications. After reading, you will have a better understanding... unavailable, try substituting with knockdown approaches using siRNA or shRNA. If this alternative is ... alternative, albeit less ideal, verification approaches discussed in the Antibodies 101: Validation post...Secondary antibody specificity  In indirect staining approaches, secondary antibody specificity must also be ...

...you with these, so keep reading! Good luck, and happy optimizing!  Recommended Reading Issa, S. S., Shaimardanova.... A. (2023). Various AAV Serotypes and Their Applications in Gene Therapy: An Overview. Cells, 12(5), ...A. (2021). Next Step in Gene Delivery: Modern Approaches and Further Perspectives of AAV Tropism Modification...AAV-Mediated Anterograde Transsynaptic Tagging: Mapping Corticocollicular Input-Defined Neural Pathways...Retrograde Serotype Viral Vectors 101: Viral Applications...

...making it very useful for genetic engineering approaches. Most importantly for this technology, however...piggyBac™ technology can be used for numerous applications, including transgenesis, gene-trap screens, ...ssODN) as the donor [6]. The problem with this approach is that, since there is no selection, you will...technology that may be used for a variety of research applications, including making transgenic cell lines and ..., while avoiding the pitfalls common to other approaches. Many thanks to our Guest Bloggers Amar Singh...Fingers, TALENs, and CRISPR-Cas9 nucleases. Mathew applies his specialized knowledge to help researchers identify...

...created by the Recipient Right to file patent applications on modifications or inventions Right to publish...can even sign electronically or choose to auto-approve MTAs. Whatever your preference, we are here to ...to make it fast and easy. Fast Approval: The UBMTA used by Addgene is internationally recognized as...transfer officers are familiar with the UBMTA and are happy to agree to it. Their confidence and familiarity...familiarity with the UBMTA result in quick approvals, so your plasmid requests can get to your lab faster. For...

...have used fluorescent proteins to observe what’s happening inside cells for at least a couple of decades....of hues. While those tools are great for many applications, Verkhusha and his lab at Albert Einstein College...exactly what they reported doing in a paper that appeared in Nature Biotechnology back in 2011 and then ...Verkhusha’s team reported another advance in the application of near-infrared fluorescent proteins, iRFP670...contrast agents for two-color imaging in animals. The approach relies on ultrasound waves as opposed to light...

... Normal Variables – Independent Variance – Approximately equal Data type - Quantitative If your data ...null hypotheses. Benjamini uses a complementary approach which identifies a p value cut off. This method...all generally yield a similar result and are appropriate to use if all sample comparison is needed, without...keep your friendly neighborhood biostatistician happy by making your statistical analysis plan when you...right statistical test, just do your homework and happy testing!   References Wissing, Dennis R., and Timm...Variables: A Monte Carlo Simulation Study. Front. Appl. Math. Stat. Volume 2 (2016). doi.org/10.3389/fams...

...increase specificity. This modular approach made ZFNs highly appealing to researchers.  TALENs In 2010, ...fiction, was also a trained biologist. This is apparent in his work The Island of Dr. Moreau (1896) where...made SciFi director Ridley Scott wonder what would happen if these corporations would extrapolate genetic...successful in the box office, indicating the insatiable appetite of the moviegoers for dinosaurs wreaking havoc...also transcended into the world of SciFi. This is apparent in the use of the word ‘CRISPR’ for anything associated...

...tag to remain on your protein for downstream applications Below, we describe a few of the most common...not to perturb protein function for downstream applications. Tag cleavage must be performed after primary...also provides a visual detection mechanism via applications such as western blot and immunofluorescence....dimerizes in solution, which can affect downstream applications. Also, purifying GST by affinity chromatography... are generally best suited for their primary application. Ready to start tagging? We are here to help ...

...beneficial for use in therapeutic or behavioral applications where drawn out effects are required. Targeted... subcellular regions of a neuron. Optogenetic approaches excel at discrete cell or tissue-subregion targeting...to the limits of light delivery. Chemogenetic approaches excel at broad tissue or system-wide targeting...chemogenetics. Note that, as with all tools, both approaches suffer from unique sets of off-target side-effects...EndogenousNot easily controlled Unique advantages Mapping the brainDissecting circuits Studying GPCR pathways...

...users have recently begun to develop CRISPR-based approaches for studying gene function in these systems. ...use in a fetal-lung derived organoid system, the approach is intended to be amenable to organoids derived...tissue-types. By bringing together CRISPR-based approaches and organoid models, toolkits like this one provide...trigger seed germination of parasitic plants, this approach was also used to build and validate a ratiometric...parasitization of cereal crops. As a result, agricultural applications of these novel biosensors could have a major...

...thermophiles, bacteria must be grown at lower temps. This approach only works for facultative thermophiles (high ... occurs? This is different than the engineered approach used to make eSpCas9 and SpCas9-HF1. By mining...SpCas9 RNP’s and a guide targeting a site with an overlapping PAM. In both cases, GeoCas9 and SpyCas had similar...could also be used with in vitro molecular bio applications that require high temperature reaction conditions...always, the plasmids are available from Addgene. Happy CRISPRing! References Harrington, Lucas, et al....