...TOPO-TA) cloning; however, the TOPO cloning technique has also be adapted for blunt end cloning. So how does...PCR products. Keep in mind that Taq polymerase has an error rate of about 1 in 3,500 bases. Typically...Resources on the Addgene Blog Perform Site Directed Mutagenesis by PCR Use REPLACR Mutatgenesis to Easily Generate...

...emotions, and even what therapeutic properties it has. As with other works of Dr. Sacks, Musicophilia includes... pretty biased as the founder, but our book club has become one of my favorite parts of Addgene’s culture...love a book series that features a young female protagonist, some clever magic, and a satisfying conclusion...

...effective than both AAV9 and AAV5, which has tropism for lung cells, and has increased overall transduction throughout...this definition: A systemic capsid is a capsid that has been engineered to have enhanced function after intravenous...both). Therefore, a high-efficiency systemic capsid has better efficacy than AAV9 and/or its intermediate...used in LY6A-nonpermissive mouse strains. **AAV1-X1 has not been tested in rats. For more information on ...neurons in the CNS (Chan et al., 2017). AAV-PHP.eB has frequently been used for screening candidate regulatory...depositing manuscript) across the brain. AAV-BI-28 has been tested in C57BL/6J mice and effectively transduces.... 4c, Jang et al., 2023; Brown et al., 2021.) It has a similar transduction rate to AAV-PHP.eB in the ...

... technique developed in David Liu’s lab in 2019, has added considerably more precision and flexibility... the canonical base editing window, base editing has higher efficiency and fewer indels than prime editing... repair (HDR) stimulated by double strand breaks has been widely used to generate precise changes. However...nicking of the unedited strand only after editing has occurred and reduces indels by 13-fold (Anzalone ... al., 2024; Yarnall et al., 2023) Prime editing has a lot of promise for treating genetic diseases. In...ones in Addgene’s repository later in the post. Advantages of prime editing Less constrained by PAM sequence...

...increase the sensitivity of detection (14-20). LDR has also been extended to multiplexed probe sets that...high-sensitivity multiplexed format (21, 27). This methodology has been used to rapidly screen buffer conditions for... 200 mM KCl at pH 8.5. This optimization method has been used internally at NEB to screen additional ...additional ligases, conditions, and formulations and has led to the development of the new HiFi Taq DNA Ligase....Numerous applications have been developed to take advantage of the high fidelity of Taq and other thermostable...

...CRISPR/Cas editing toolbox for C. elegans Recent work has further improved the efficiency of genome editing... (Farboud et al., 2019). Work from the Mello lab has dramatically improved RNP editing with linear repair...elegans (Ebbing et al., 2017). Because C. elegans has an AT-rich genome, it can be difficult to find an...whereas import of CRISPR technology into C. elegans has been a transformative innovation for our community...-CRISPR and co-conversion approaches have the advantage of being used in any genetic background, but require...

...infect dividing and non-dividing cells.  Addgene has an extensive collection of lentiviral plasmids created... immortalized with adenoviral protein E1A, which has been shown to repress expression of HIV-1 LTRs (more...the deletion reduces the risk of insertional mutagenesis by limiting the interaction between viral LTR...potential for oncogenesis through insertional mutagenesis. These risks are largely based upon the vector...

...the post) has a number of helpful suggestions.  Gels Gel issues usually come down to voltage and time....ladder. Unstained ladders contain either ‌a protein tag or an IgG binding site, allowing you to visualize...bands, your voltage is likely too high. If you’re seeing a smiley face at an appropriate voltage (usually...can happen with large proteins, try lowering the voltage and using a longer run time.   Samples can run ...packs can help keep the system cool, as can lower voltages.  If your small protein is over-transferring, ...transfer Increase transfer time Increase voltage Increase SDS/decrease alcohol in transfer buffer...Psi membrane Semi-dry transfer Decrease voltage Decrease transfer time Decrease SDS/increase...

... Addgene’s people, culture, and workplace. Rumor has it that a few Addgenies* may even be open to dancing...Social Media Are you following us on Twitter or Instagram (or Facebook or LinkedIn) but want to hear and...love sunshine and warm weather - check out our Instagram all summer long to see what kind of shenanigans...

...bind DNA targets without changing them. Thus, it has been widely adapted for a wide variety of applications...inhibition, among others. Over the past few years, dCas9 has also become a robust visualization tool to study ...guide sgRNA-Sirius-8XMS2 which will recruit the HaloTag with red fluorescent ligands to the loci of interest...

...supernatant during the production phase. However, this has not been well characterized as it is not a common...microvesicles as a vehicle for drug delivery that has recently gained interest special contexts, such as...researchers discovered that these particles have some advantages over the conventionally purified AAV vectors....

...HiFi DNA Assembly method from New England Biolabs® has minimal upfront requirements and allows you to expediently...yeast that same day.  The field of synthetic biology has led to advances in biofuel generation, green chemistry...HiFi can simplify multi-position site-directed mutagenesis by assembling multiple fragments containing the...

...the indicated function. One huge reason CRISPR has become such a popular genome editing tool is its developers...next-generation sequencing to ensure that the library has retained gRNA representation, produce high titer ...that particular organism available at Addgene. Percentages represent the fraction of that total with the...

...and the master plasmid in one step. The kit also has other unique aspects that make it especially well-suited...for CRISPR/Cas9 (also available from Addgene) only has two plasmids. In many cases, CRISPR may be more efficient...together, there are occasional errors. That can be advantageous if you are trying to knock out a gene. One can...

...Cytotoxicity can also be cell type dependent: if a cell has few receptors for a specific envelope protein, it...lentiviral vector to be transported retrograde and also has higher neuronal transduction efficiency and a better...different viral membranes, tropism, and advantages and disadvantages: Table 1 (Joglekar et al., 2017)  Table...its own, it needs to be pseudotyped. This takes advantage of the EnvA-TVA receptor-envelope system (Wickersham...

...mammalian cells. Each of these systems has their advantages and disadvantages. Briefly, bacterial and yeast cells...recombinant antibodies (rAbs) exist (though Addgene has a great collection of ready-to-use rAbs and rAb plasmids...

...purify DNA, but rather a glass powder slurry that has a milk-like appearance. To make DNA stick to the ...chaotropic salt sodium iodine (NaI) solution. NaI has two purposes in this protocol: 1) it solubilizes ...DNA without a kit. DNA purification kits have advantages: they are convenient and provide uniform, consistent...least entraps nucleic acids. The Botella lab took advantage of this property and developed a protocol that...

...therapeutics to treat neurodegenerative diseases. He has earned a B.S. in microbiology and subsequently obtained...Newland is General Counsel at Atalanta, an early-stage biotech company that is developing RNAi therapeutics...this past year and a half. My wife and I went to Patagonia in February of 2020 and came back just a week ...

... and microbe cookies! 🤗 🤗 🤗 We hope everyone has a safe and festive Holiday season. 🙌 🥂 🎄 🎉 😷...showcase it all!           View this post on Instagram                       A post shared...dyed with real microbial pigment. Check out the hashtag to see the description of all the ornaments. #DeckTheLab...

...cornerstone for The Addictive Brain. Our community has also requested live question and answer sessions ...sessions led by scientists, which is hosted on our Instagram channel every Sunday. What's planned for the upcoming...participate in our one-hour Live Q&A session on Instagram every Sunday. Undergraduate and graduate students...background. Social media is a powerful tool. Use hashtags effectively. This would help you direct your content...