...packaging capacity is limited to ~4.7kb, or roughly half the packaging limits of lentiviral and adenoviral...multiple copies of both vectors. Examples of use: Halbert et al., 2002 used an overlapping vector with 440...heterodimer, then splicing of the pre-mRNA of the two halves of the transgene will reconstitute the full-length...A of the transgene contains the promoter, the 5’ half of the transgene, a splice donor sequence and the... of homology, a splice acceptor sequence, the 3’ half of the transgene. The full length gene is reconstituted...124–130 . https://doi.org/10.1038/sj.mt.6300322 Halbert CL, Allen JM, Miller AD (2002) Efficient mouse ...

... Awards: Melissa Chiasson, Elizabeth Unger, and Michal Bernach. Melissa Chiasson: Quantifying the abundance... in vitro against different drugs and dosages.  Michal Bernach: Developing an artificial leaf replica ...plant-bacteria and bacteria-bacteria interactions  Michal Bernach is a graduate student in Mitja Remus-Emsermann...surfaces. His system of choice is the Arabidopsis thaliana - Pantoea agglomerans interaction. But the shape...bacteria. To compare the artificial leaf to A. thaliana leaf, Bernach plans to use a suite of fluorescent...

...algorithms, such as SIFT or PolyPhen to approach this challenge, using attributes of cancer variants such as evolutionary...to making cancer precision medicine a reality. Challenges for cancer variant characterization For many ...validation experiments. But, there have been two huge challenges preventing us from testing this hypothesis: ...Accelerator Plasmid Collections To overcome this first challenge, we’ve developed the Broad Institute Target Accelerator...cancer variants To overcome the second major challenge, we developed assays to test large numbers of ...

...SNAP-tag, and HaloTag fusions, while the anti-HA frankenbodies include mEGFP, mCherry, and HaloTag. ...screening technique, researchers identified a synthetic lethal interaction between MARCH5 and WSB2 genes using...fusion proteins, such as Cas9 and Cas12a, creates challenges for in vivo delivery and cell engineering applications..., is composed of only 529 amino acids, less than half the size of either Cas9 or Cas12a. Despite its small...

...optogenetics.  Challenges of engineering channelrhodopsins There are three main challenges to engineering...overcome channelrhodopsin design challenges To overcome these challenges, the Arnold and Gradinaru Labs ...lower multiplicity of infection (MOI) makes it challenging for ChR2(H134R) to function because it’s lower...

...since the Future of Research Symposia Ongoing challenges facing the scientific enterprise and the way ...landscape of the scientific enterprise will provide challenges and opportunities for us all.  For example, implementation...categorize all postdocs at institutions, will be challenging. At the same time, there is much opportunity ...the pressure on for further increases will be challenging, but will be aided by the development and implementation... is an important way to push the balance from challenge to opportunity. We hope to see you at the upcoming...

...proteins to form the Cascade targeting complex is a hallmark of Type I CRISPR-Cas systems, in contrast to single-protein...with the transposition protein TniQ for targeting (Halpin-Healy et al., 2020). Genomic targets require an...creates a 5-bp target-site duplication (TSD), a hallmark characteristic of the Tn7-family of transposons...follow him on twitter @Vopinator.     References: Halpin-Healy TS, Klompe SE, Sternberg SH, Fernández IS...doi.org/10.1038/s41586-019-1849-0 Klompe SE, Vo PLH, Halpin-Healy TS, Sternberg SH (2019) Transposon-encoded...

...solutions to complex challenges. As a cross-disciplinary researcher, the challenge of keeping up on studies... Sharing your research with the world can be challenging. After months, or years, of grueling effort to...biology or international relations.  Many of the challenges in science today are too complex for any single...making publishing their research all the more challenging even when it is of quality). On the site you ...

...externally-timed bands of color with HaloTag-iPAK4 by applying different HaloTag dyes and showed they could record...local proteome, but some applications have been challenging or inaccessible. The Ting lab recently made two.... B) HEK cell with an iPAK4 fiber labeled with HaloTag-iPAK4. Scale bar 10 µm. C) Images of iPAK4 fibers...fibers in cultured rat hippocampal neurons. HaloTag dyes are switched from yellow to red at 0 h and back...

...DSB) are lethal. Addgene depositor Luciano Marraffini’s lab took advantage of this lethality to design...organisms, CRISPR-induced double stranded breaks are lethal because the non-homologous end-joining (NHEJ) repair...Cas9 will cleave the gene of interest, inducing a lethal DSB.     This system is distinct from those used...While the activation of transcription is a more challenging problem than repression, scientists have deposited...Streptomyces, yet genome engineering remains challenging for most organisms in the genus. Work from the...

...biomedical model organism.  Arabidopsis thaliana Arabidopsis thaliana is one of, if not the most, recognized...Arabidopsis genetics research development. Arabidopsis thaliana was fully sequenced by the year 2000, becoming...properties affecting adaptations.  Arabidopsis thaliana flowers. Image from BlueRidgeKitties. If you...functions in an ecological context with Arabidopsis thaliana. ELife, 4. https://doi.org/10.7554/elife.06100...

...be the way to accomplish this tough job! Kamel Khalili’s lab at Temple University has demonstrated two ... a well-established viral reservoir. A second challenge for CRISPR HIV-1 therapies is sequence specificity..., Kaminski R, Fagan PR, Putatunda R, Young WB, Khalili K, Hu W. CRISPR/gRNA-directed synergistic activation...T, Tedaldi E, Napoli A, Zhang Y, Karn J, Hu W, Khalili K. Elimination of HIV-1 Genomes from Human T-lymphoid...PMID: 26035283. PubMed Central PMCID: PMC4649911. Khalili K, Kaminski R, Gordon J, Cosentino L, Hu W. Genome...

...Precision genome editing faces the challenges above and more. One chief challenge is upregulating HDR in mature...complex oligonucleotide treatment comes with many challenges, researchers have begun to explore genome editing... the time to clinical approval. Even with the challenges precision editing faces, it’s encouraging to ...WX, Scott DA, Gootenberg JS, Kriz AJ, Zetsche B, Shalem O, Wu X, Makarova KS, Koonin EV, Sharp PA, Zhang...

... demethylase to demethylate enhancer regions (Mendenhall et al., 2013). By comparing gene activation when...sorting and selection of transduced cells. Grant Challen’s lab also created constitutive (pCMV-dCas9-D3A)...contributed to updating this article. References Haldeman JM, Conway AE, Arlotto ME, et al (2018) Creation...Fowler T, Rees R, Kramer A, Martens A, Edwards JR, Challen GA (2016) Reprogrammable CRISPR/Cas9-based system...Find plasmids from this publication at Addgene. Mendenhall EM, Williamson KE, Reyon D, Zou JY, Ram O, Joung...

...information. Many of our technology platforms like HaloTag® Fusion Protein, codon-optimized Firefly luciferase...concentrations of biological molecules. FRET sensors face challenges of photobleaching, autofluorescence, and, in ...cyan-excitable donors, phototoxicity. Another challenge to using FRET sensors comes when employing optogenetic...Researchers have sought to alleviate many of these challenges by exchanging the fluorescent donor for a bioluminescent... analysis products at Promega.     References 1. Hall, M.P., et al. (2012) Engineered luciferase reporter...

...interchangeable labeling using the HaloTag® system, the new Janelia Fluor HaloTag Ligands expand the self-labeling... self-labeling proteins would be SNAP-tag® and HaloTag®. These consist of a genetically encoded enzyme...Hayashi-Takanaka et al., 2014). Other dyes may prove challenging to stain intracellular compartments in in vivo...2017). These dyes were adapted into ligands for HaloTag® and are currently offered at Promega. Due to the...

...and represent the diversity of tools we share on behalf of scientists. 9. Plasmids 101: Positive and Negative...fluorescent proteins at Addgene. 7. Designing Your Chalk Talk for the Academic Job Interview by Erik Snapp...blogger Erik Snapp breaks down how to prepare a chalk talk for an academic job interview including defining... research vision, organizing your vision into a chalk talk, and tips for practicing.  6. The Fluorescent...

...annurev-neuro-111020-100834 Challis, R.C., Ravindra Kumar, S., Chan, K.Y., Challis, C., Beadle, K., Jang, ...or lumbar spine. Image and caption courtesy of Challis et al., 2022. Where do they come from? Systemic...https://clover.caltech.edu. Figure adapted from Challis et al. 2022. Image created with BioRender.com. ...low doses a regional bias towards the cortex and thalamus emerges. AAV-PHP.eC effectively transduces neurons... Resources and references Recommended reading Challis R.C., Ravindra Kumar S., Chen X., Goertsen D., ...https://doi.org/10.1016/j.crneur.2023.100086. Challis, R. C., Ravindra Kumar, S., Chen, X., Goertsen,...

...promoting domain. Each half was fused with one of the LOVTRAP domains. When both halves of the modified EB1...growth and extension. Now, they split EB1 into two halves (Fig. 1): one containing the microtubule binding...expansion of the genome editing toolkit. However, a challenge in optimizing base editing technologies across...

...antibody selectivity. Additionally, it can be a real challenge to troubleshoot RNAi failures. Potential explanations...cell survival or proliferation, an additional challenge is that only partial or ephemeral knockdown will...confirmation of antibody selectivity even more challenging. Correlation with RNA or Proteomic data from ...supportive of their specificity for the protein. A challenge to this approach is that the identity of the epitope...the specific sample and protocol. This can be a challenge to confirm. As such, the approach used may need...