...bacteria, transposons can move antibiotic resistance genes between plasmids or from plasmids to the bacteria... and mechanism of replication. They contain two genes: gag and pol. The pol polyprotein encodes the reverse...transposon in vertebrates. It was discovered in Japanese medaka fish because its insertion into the fish... how they can be used in the lab. Transposon mutagenesis screens Transposons, by their nature, are mutagenic... elements, which makes them a great tool for mutagenesis screens that detect loss-of-function or gain-... these screens, the transposons encode reporter genes, mutagenic cassettes, or barcodes. When transposons...to consider when selecting a transposon for a mutagenesis screen. For example, piggyBac and Tol2 are best...

...perturbations that may play significant roles in cancer pathogenesis. Their work culminated in the first functional...help cancer researchers studying an assortment of genes. Using patient data to identify mutations of unknown...mutations with strong phenotypic effects can mask weaker ones. Thus, the need for a faster and higher-throughput... team identified 1,049 perturbations in several genes spanning 33 cancer types. The types of mutations...lentiviral constructs using the High-Throughput Mutagenesis and Molecular Barcoding (HiTMMoB) technique (.... Plasmids carrying the corresponding wild-type genes were constructed as well. They called this library.../F3 and MCF10A are growth factor dependent cell lines and any mutations that that confer proliferation...

...Syndrome (a disease that results in combined deafness and blindness) is caused by defects in the gene MYO7a...Jean Bennett started working on ways to deliver genes to correct genetic conditions over 25 years ago ...to the identification of several disease-causing genes, including the RPE65 gene responsible for an autosomal...autosomal recessive, severe, childhood-onset form of blindness name Leber’s Congenital Amaurosis (LCA). Shortly...treatment, famously visited Congress to help increase awareness about the potential of gene therapy!). It would... packaging capacity is well suited for larger transgenes. Unfortunately it has limited tropism in the ...used in clinical trials, a variety of engineered ones have shown great promise in preclinical studies ...

.... Maintaining openness and looseness of spirit The second crucial element in harnessing the power of serendipity...beyond its confines. The beauty of serendipity often emerges at the intersections of disciplines. Cast your... serendipity is maintaining an openness and looseness of spirit. Amidst the rigors of PhD studies and ...routine. It encourages you to think beyond the confines of a predefined path, opening doors to novel ideas...expected. By widening your search and maintaining an openness to unexpected possibilities, you not only invite...

...Addgene also has ready-made entry clones available for many popular genes, including Hs.KRAS4a. Use our website...selection Used to clone attB-flanked genes of interest to generate entry clones Entry vector attL sites for...competent E. coli cells and select the positive clones. The entry clone and destination vector carry different... recombination step. Addgene’s ready-made entry clones can be used with a large variety of plasmids. Speed...for recombination Used to generate entry clones by TOPO cloning or by Restriction Cloning Destination...the gene of interest in the appropriate system Recombines with the entry clone to generate an expression...plasmids with Addgene. These can be used to express genes in a variety of model organisms. Use the links below...

... the paucity of Cre-dependent mouse lines for overexpressing genes within discrete cell populations makes...libraries. However, using CRISPRa to target individual genes can be relatively laborious because multiple plasmids...several sgRNAs must be screened to identify effective ones. Simplified in vivo delivery using SPH mice CRISPRa...neurons over other promoters, including ubiquitous ones such as EF-1α and CAG. Figure 2: Dual lentivirus...resulted in hundreds of differentially expressed genes between Bdnf I-overexpressing neurons and Bdnf IV-overexpressing...IV-overexpressing neurons. Among the upregulated genes common between the Bdnf I and Bdnf IV overexpression...overexpression variants were other immediate-early genes associated with neuronal activity, such as Fos, Egr3,...

... for a molecular biologist who wants to express genes from one organism in another? Let’s take a look ...correlated with codon bias across all endogenous genes in E. coli and S. cerevisiae (Tuller et. al., 2010...usage frequency in a reference of highly expressed genes (Sharp and Li, 1987). In 2000 the Ikemura lab created...an electronic dataset for codon-usage of 257,468 genes across 8,792 organisms (Nakamura et al., 2000).  ...2017. Most commonly, scientists codon optimize genes from eukaryotic organisms for expression in prokaryotic...optimizing a human gene for expression in E. coli. Genes are also optimized for expression in mammalian cells... lab optimized the C. elegans GluCl ion channel genes for expression in mammalian cells. This was the ...

...CRISPR-Cas can harness the power of MoClo to expand both targetability and effectiveness. Perhaps the most...and Plant Parts Kit III (Gantner et al., 2018; Chamness et al., 2023). The Plant Parts II kit has a focus...for both monocots and dicots (Hahn et al., 2020; Chamness et al., 2023; Stuttmann et al., 2021; Grützner...CRISPR components expressed from different plasmid backbones. They noted that backbone differences such as ...Biotechnology, vol. 17, no. 1, Springer Science and Business Media LLC, 13 Nov. 2017. Crossref, https://doi.org...Crossref, https://doi.org/10.1021/sb4001504. Gantner, Johannes, et al. “Peripheral Infrastructure Vectors and... https://doi.org/10.1371/journal.pone.0197185. Chamness, James C., et al. “An Extensible Vector Toolkit...

...experiment. Nucleic acid based machines Recombinase-based State Machines Enable Order-dependent Logic in... cell. These so-called recombinase-based state machines could potentially be used to report upon and/or...post to learn more about recombinase-based state machines and how they function. Adapting Toehold Switches...replication and transcription as well as reporter genes. To learn how you can use minigenomes in your research... describes R-bodies, “force generating protein machines that can extend and retract over many microns ...also showed that R-bodies can rupture E. coli membranes. Read the full post for potential applications...Adeno-associated viruses (AAVs) are commonly used to deliver a genes of interest to a variety of cell types in-vivo. ...

...: rare. “Nature has many genes where there are not rare codons and many genes where there are instances...trend of sourcing genes from distantly related organisms. In 2006, less than 1% of the genes deposited at ...synthetic or natural with 97.7% accuracy. Classifying genes as natural or synthetic But how can you tell if ...in a test set of 173 known synthetic and natural genes.  In designing and optimizing their algorithm, ...synthetic. But nine years later, over 20% of the genes deposited that year were synthetic. They also found... Proteobacteria - the most frequent transfer of genes is from Proteobacteria to mammalian expression vectors...Thompson. "Gene synthesis allows biologists to source genes from farther away in the tree of life." Nature communications...

...unstructured peptide and therefore consisted of glycines, serines and other small, non-aliphatic amino acids...fluorescent proteins to oligomerize is known as “stickiness.” Since the fluorescent tag should operate as... per unit.  There are several ways to measure stickiness. In vitro, ultracentrifugation or gel filtration...oligomerization in cells. In addition to measuring stickiness, several researchers have also collected data...fluorescent protein characteristics such as the brightness, photostability, maturation time, and acid sensitivity...characteristics are important, but have less priority than stickiness. In my experience, fluorescent proteins that ...mediated HDR) enables the tagging of endogenous genes close to native expression levels. However, it is...

...consider effective brightness of NIR FPs in cells, in addition to their molecular brightness (a product of ...Molecular brightness vs. iRFP713, % Oligomeric state Photo-stabilityc, t1/2, s pKa Brightness in HeLa ...coefficient) (Shcherbakova et al., 2015). The effective brightness reflects the concentration of BV-containing molecules... photoreceptors have higher apparent cellular brightness than previously reported widely used dimeric ...not yield to (m)iRFPs in cellular and molecular brightness (Table 1).  NIR reporters and biosensors NIR ...-the-art NIR FPs, which combine high cellular brightness and monomeric state, are highlighted in blue....Oliinyk, O.S. & Verkhusha, V.V. How to Increase Brightness of Near-Infrared Fluorescent Proteins in Mammalian...

...the secretory pathway. Those can include cytokines and chemokines; enzymes like kinases, phosphatases, and...interchangeably to refer to any technique that combines a marker with a signal. For clarity, Addgene uses...of such targets include transcription factors, histones, and DNA repair enzymes. Note that when using ...label cytosolic proteins alongside the intranuclear ones. You do not need to use a separate fix/perm cytosolic...non-antibody-based stains that bind DNA or free amines of proteins. Depending on the application, they...membrane-impermeant, and the cells of interest can be the ones that are positive or negative for the dye. The main... can only enter cells with compromised plasma membranes — i.e., dead cells — while live cells are protected...

...lab that expresses the HIV gag, pol, rev, and tet genes. It can be used with 2nd or 3rd generation lentiviral...the Zhang lab expresses Cas9 from Streptococcus pyogenes with an invariant sgRNA scaffold and cloning sites...Addgene!) pX330-U6-Chimeric_BB-CBh-hSpCas9 - The S. pyogenes Cas9 (SpCas9) has been widely popular in genome...genome engineering. For use in human cell lines, the Zhang lab codon-optimized SpCas9. Using the BbsI restriction...cassette that makes it less efficient in some cell lines. pSpCas9(BB)-2A-Puro (PX459) V2.0 was made available...from the Trono lab that contains the gag and pol genes. It also requires pRSV-Rev and the envelope expressing...lentiGuide-Puro - This plasmid expresses the S. pyogenes CRISPR chimeric RNA element with customizable ...

...between MARCH5 and WSB2 genes using Cas12a’s ability to knock out multiple genes in parallel. Lastly, this...individually (~20,000 lanes/50 mLs) or through two polycistronic expression vectors (3570 lanes/50 mLs) coexpressing...frankenbodies to amplify the signal.  Liu et al. Genes Cells. 2021. https://doi.org/10.1111/gtc.12893, ...screening methods have primarily used Cas9 to knock out genes, researchers at the Genetic Pertubation Platform...library. Overall, Cas12a’s ability to target multiple genes within a single cassette provides increased simplicity...individually expressed proteins run in separate lanes. B: individually expressed and coexpressed proteins...gene activation for both reporter and endogenous genes. It is highly specific in mammalian cells without...

...Research the brightness of your fluorophores before buying and try to inversely pair brightness with abundance...sensitive moment.   Fig 1: Blugene fading into darkness because they accidentally left their antibodies...Type Westerns and ELISAs mostly use HRP, a chemiluminescent conjugate that can be activated to emit light...we'll keep it simple. If you follow the two basic guidelines below, you’ll likely be able to create a small...small, usable panel.  First, fluorophores vary in brightness. You don’t want a super-bright fluorophore, like...good match for your experiments, and avoid the awkwardness of getting the right antibody with the wrong ... Boston University Flow Core. Fluorochromes Brightness Chart. Boston University Flow Core. Retrieved...

...achieve success in this business. A good presentation can help win a business plan competition and advance...LabSpaces Blog Find a Buisness Plan Competition Good Relations - PR for the Biotech Business - Posted on Nature.com... Dr. Mike Koeris, co-founder and current VP of Business Development & Operations of Sample6, for advice...2009 and tested in the “market” by competing in business plan competitions. After much learning, the focus... investors to the next stage of diligence." In business competitions everything is conceptual, and the...focusing on concept development and participating in business competitions. When the team was unable to secure...became first COO, then moved on to become VP of Business Development & Operations. Starting a biotech...

... and release IL-36 cytokines. Keratinocytes are the main source of IL-36 cytokines in the skin making ...surrounding IL-36 biology. “Little is known how IL-36 cytokines actually get out of keratinocytes and induce their...researcher at University Hospital Zurich. IL-36 cytokines are a potent inducer of an autoinflammatory loop...Congratulations Mark Mellett! Siddharth Prakash: X chromosome genes involved in Turner syndrome Turner syndrome (TS)...can lead to cardiac developmental defects. Eight genes from the X chromosome are expressed at significantly...from Addgene, the lab will express these candidate genes in TS iPSCs to identify whether any rescue cell ...CRISPR plasmids to generate deletions of candidate genes in euploid iPSCs. “The BeHEARD Addgene reagents ...

...variety of genes with GFP. These plasmids are available through Addgene, and the stem cell lines we’ve made...is still inefficient, particularly in human cell lines. Furthermore, the quality control necessary to ensure...information and images generated from these cell lines can be found in the Cell Catalog on our data portal...different human cell types, and while most of the genes we’ve tagged are widely recognized markers of cellular...initial set of 10 plasmids representing 10 human genes and will add more to the collection later this year...everywhere To find out more about gene edited hiPS cell lines generated using the Allen Institute for Cell Science...collection includes systems for tagging mammalian cell lines, Drosophila cells, and C. elegans. In addition to...

...fluorescent proteins, and luminescent proteins. While fluorescent proteins and luminescence have been widely used...maturation times, and fitness cost of expression. They found that the cellular fitness cost varied greatly...plasmid that combines the most desirable features: intense color, fast maturation, and low fitness cost yet...striking visuals from fluorescent jellyfish, bioluminescent phytoplankton, and colorful corals, to name...instrument-free detection. Unlike fluorescence or luminescence, which require UV lamps, fluorometers, or luminometers...3: pTOX plasmids expressing amilCP and tsPurple genes. Image: Lazarus et al., 2019. One such lab is...

...the plant hormones strigolactones By Fabienne Furt Current methods used to detect strigolactones (SLs) are...editing in organoid Biosensors for detecting strigolactones The CRISPR Corner New ready-to-use viral preps...et al. 2021. The Organoid EasyTag workflow combines thoughtfully designed plasmid components with ...investigating genetic mechanisms related to morphogenesis, cell fate specification, and human disease....library from Alec Kimmelman targets mouse metabolic genes with ~6 gRNAs per gene.  CRISPR-Act3.0, developed...from Xiaole Shirley Liu's lab targets all mouse genes with 10 gRNAs per gene. The guides are optimized...

...inhibit, activate, or knockout many genes at a time to identify ‘hits’ - genes that affect disease progression...progression or onset. These hits can reveal individual genes or sometimes even genetic pathways that are worth...exist a drug which targets some of the candidate genes! Disease model generation CRISPR has made the generation...both individual and combinatorial drugs on for effectiveness prior to patient treatment. Target validation... revolutionized the generation of knockout cell lines which has made this control step extremely accessible...your drug(s). If a drug is equally toxic to both lines, it probably has either high toxicity or high off...and Resources References Fellmann, C., et al., Cornerstones of CRISPR-Cas in drug development and therapy...

...neurotransmission. But with approximately 950 different human genes encoding GPCRs (Takeda S et al., 2002) and many ...can be measured using a technique known as Bioluminescence Resonance Energy Transfer (BRET). BRET is similar...BRET is that the energy transfer is between a luminescent donor and a fluorescent acceptor. TRUPATH uses...measures heterotrimeric G protein dissociation by bioluminescence resonance energy transfer 2, or BRET2. With... constructs. Cells are then treated with the luminescent enzyme substrate coelenterazine, followed by ...endogenous ligand or drug. In the inactive state, the luminescent chimeric protein Gα-Rluc8 and Gγ-GFP2 will be...transducerome on a single GPCR, showing the effectiveness of using TRUPATH as a screening technique. Lastly...

...deletion of one or several of these genes permits the expression of genes farther downstream in the construct...Brainbow uses a Cre/lox system to label neurons. Genes encoding the fluorescent proteins are organized ...developed, which differ in the number, stability, and brightness of the fluorescent proteins used in the system... which uses Cre to trigger the inversion of two genes aligned head-to-head. Another variation, Brainbow... to trigger the inversion of two tandem sets of genes aligned head-to-head and the deletion of one of ...located directly upstream of the fluorescent protein genes, Cre recombination is required before fluorescence...brainbow,” is a collection of transgenic zebrafish lines that allow for multicolor labeling of an entire ...

...during and after embryogenesis. Addgene has over 600 plasmids that contain Xenopus genes and a tool-kit ... concluded that nearly all disease-linked human genes have counterparts in the rat (Gibbs, et al., 2004...collection of plasmids containing Rattus norvegicus genes! Mexican axolotl (Ambystoma mexicanum) Ambystoma...entire axolotl genome, revealing an abundance of genes related to limb regeneration (Vieira et al., 2019...for use in CRISPR and Tol2 transposase induced transgenesis.  Xenopus Xenopus, or frogs, are one of the ... for scientists to study human disease-relevant genes in such a controlled environment displays how essential...molecular tools, such as gene manipulation and mutagenesis techniques, are available to the scientific community...

...the location of genes. With this knowledge, researchers were able to target specific genes for deletion ... at Harvard University. Just as computers, cell phones, and cars become more technologically advanced ...fluorescently labeled probes. Colorimetric and luminescence based detection methods are also commonly used...ddNTP, and then run these reactions out in separate lanes of a gel. Since the sequencing gel will separate...nucleotides were replaced with fluorescently labeled ones allowing for fluorometric-based detection. This ...generally considered first generation DNA sequencing machines (9). Today, similar technologies, sometimes referred...make techniques used to generate transgenic mouse lines, such as BAC transgenic or traditional knockins/...

... our purposes? In search of simple biological machines R bodies can repeatedly extend and contract...using R bodies: simple, force-generating protein machines that can extend and retract over many microns ...with proteins and remain functional, 2) break membranes outside of their natural context or 3) be modified...demonstrates that R bodies can function to break membranes of different organisms. And finally, we demonstrated...demonstrated that R bodies can be “tuned” with mutagenesis to extend at different pH values. Using a screen...in a particular region of the R-body. Further mutatgenesis of this region produced R-bodies that could ...could use them to lyse cells (and surrounding membranes) in keeping with their natural function (see below...

... disease, cholesterol gallstone disease, senescent illnesses, and the metabolic syndrome. Coregulators...functional groups of genes involved in the execution of a specific regulatory regime (such as genes involved in...ligand-dependent sensors for a diverse set of fat-soluble hormones, vitamins, and dietary lipids. Included in this...remain unknown. Together, NRs govern expression of genes involved in a broad range of reproductive, developmental...For example, the phosphorylation status of SRC-3 defines its association with other members of the complex...as in important physiological states such as adipogenesis and macrophage activation. Our mass spectrometry-affinity...

...2014). Microbial rhodopsins are encoded by opsin genes, and include bacteriorhodopsins, halorhodopsins,...control the flow of electric charges across cell membranes and maintain membrane potential in response to...light-sensitive, so the selective expression of opsin genes in targeted cells makes it possible to control neuronal..., 2014). This could be overcome by fusing opsin genes to cell-type specific promoters; however, those ...those usually drive weak expression of downstream genes. Strong cell-specific promoters, on the other side... circuitry behind the mechanisms of sleep and anesthesia. In Cre transgenic mice, they targeted the dopamine... previously subjected to steady-state general anesthesia (Taylor et al., 2016). These very same neurons...

.... These could be genes, specific polymorphisms, including antibiotic resistance genes. This approach successfully...Genome editing tools for bacteria CRMAGE system combines CRISPR and recombineering-based MAGE The Nielsen...CRMAGE system is a fast, multiplexable method that combines CRISPR and recombineering-based MAGE (Multiplex...Yang’s lab describes a two-plasmid system that combines recombineering with CRISPR to create a system ...effectively silenced transcription of one or more genes in E. coli. and in a mammalian cell line (Qi et ... repressed or activated transcription of select genes in E. coli. Furthermore, to demonstrate wider applicability... the Husson lab used CRISPRi to study essential genes in M. tuberculosis. dCas9 and a gRNA are expressed...

... AdVs are stripped of regulatory genes E1 and E3. Without these genes, AdVs cannot replicate on their ...produced at high titers, can accommodate large transgenes, transduce quiescent and dividing cells, and ..., or high-capacity (HC-Ad__ are devoid of viral genes and therefore can accommodate up to 36 kb. Gutless...can be produced in E1-containing mammalian cell lines such as HEK293 cells. 1st generation AdV cloning...immunogenicity. In addition to E1/E3, non-structural genes E2 and E4 are deleted in 2nd generation AdVs, thereby...cloned) and pAdEasy™ which contains the adenoviral genes necessary for virus production. For detailed instructions...therapies in the new era of personalized medicine." Genes & diseases 4.2 (2017): 43-63. PubMed PMID: 28944281...

... and bleed dates and even clonal hybridoma cell lines can produce more than one mAb (Bradbury et al., ...Hybridomas can lose gene expression of the mAb-encoding genes or fail to revive after cryopreservation (Bradbury...antibodies that are generated in vitro using synthetic genes that are typically expressed from a plasmid or from... integrated sequence in a stable cell line. The genes encode the heavy chain and light chain for the antibody...Figure 2: (Left) The light chain and heavy chain genes of the antibody are encoded in a plasmid. (Right...better and better at identifying and cloning the genes for antibody expression. If starting from a protein...sequences of the antibodies and then synthesize the genes that encode those amino acids (Tran et al., 2016...

...Specially constructed plasmids can be used to replace genes in yeast, mice, or Drosophila through homologous...techniques and the plasmids used to inactivate specific genes in mammalian cells. Despite the popularity of CRISPR-based... Neomycin, puromycin, and hygromycin resistance genes are commonly used for positive selection. While ...methods have been used to create many knockout cell lines and animal models, their efficiency is very low,...easier to use and are more efficient at inactivating genes. CRISPR can target a genomic sequence and create...perfected, the production of cells and mouse knockout lines should increase. New genome engineering tools also...have tools for creating knockout animals or cell lines, please consider sharing them by depositing with...

... Cells Transformed cell lines Transformed cell lines Transformed cell lines Cargo Nucleic acids or ...cells or a selection marker to generate stable cell lines. Transient or stable expression of Cas9 and/or gRNA...contains a selection marker to generate stable cell lines. Packaging and Envelope plasmids provide the necessary...and/or gRNA in a wide variety of mammalian cell lines. Useful for difficult to transfect cell types and... cell.   Delivery methods for mammalian cell lines As mentioned above, the expression system you choose...Delivery of these components into mammalian cell lines is quite broad and includes several different methods...

...PA-FPs by performing several rounds of random mutagenesis screening for enhanced RFP variants. PA-mCherry... Color Contrast (Fold Change in Fluorescence) Brightness Excitation/ Emission (nm) PA-GFP Monomer UV-Violet...negative” photoswitching. In 2008, scientists used mutagenesis screens to find a version of Dronpa that doesn't...background fluorescence, but their single molecule brightness and ability to switch from a dark state to a ... Color Contrast (Fold Change in Fluorescence) Brightness Excitation/ Emission (nm) Dronpa Monomer UV-Violet...there are several parameters to consider, such as brightness, photostability, pH stability, chromophore maturation... is the contrast ratio- a comparison of PA-FP brightness before and after photoactivation. A high contrast...

...binding. Formaldehyde cross-links the protein (red lines), which maintains structural epitopes (orange region...proteins that need a little help crossing cell membranes as they are not able to diffuse across natively...Permeabilization essentially punches holes in the cell membranes of your sample to allow antibodies better access...example, saponin interacts with cholesterol in cell membranes and leaves many membrane-associated proteins in...detergents that interact with lipids and proteins in membranes and permeabilize non-selectively. This non-selectivity...helpful when trying to pass tough-to-permeabilize membranes and these detergents avoid some of the negative...but it is better at permeabilizing organellar membranes. Created with BioRender.   How do I choose...

...substantially enhance the brightness and photostability of the classic rhodamines and their analogs without... a greater burden on the photon budget, which determines how long and precise we can track individual ...selectivity of the fluorophore attachment, the brightness of the resulting conjugate, and the complexity... classic small molecule fluorophores, such as rhodamines (Figure 2a), were discovered in the 19th century...due to its established chemistry, small size, brightness, and cell permeability. In past decades, further... the Alexa Fluor and ATTO dyes, with improved brightness, photostability, and spectral range. Nevertheless...PMCID: PMC4344395. 3. Lavis, Luke D., and Ronald T. Raines. "Bright ideas for chemical biology."ACS chemical...

...PHP.B.  A Ly6a variant is linked to PHP.B CNS permissiveness Both the Deverman and Wilson labs determined...as non-permissive to PHP.B CNS transduction. Two genes were associated with the PHP.B permissive phenotype... suggesting it was a likely player in PHP.B permissiveness. The Wilson Lab also looked for the gene variant...mice to map the inheritance pattern of PHP.B permissiveness. The first generation of outbred mice (F1) ...~4.5 Mbp region of the mouse chromosome. Of the genes located in this stretch of DNA, Ly6a seemed the ...cells: galactose and the AAV receptor (AAVR). Cell lines lacking galactose were still transduced by PHP.B...to the CNS. The bottom line of viral vector permissiveness While PHP.B has impressive CNS transduction...

...Toth) These wasps are like the ones that nest on your house. Their nests are open so you can see them and... human interaction for all of us studying human genes in test tubes and gels). As a freshman in college... I studied development and evolution of histone genes in Strongylocentrotus purpuratus, the spiny purple...silencing in the wild. They also return to their nests so can be studied over time. They recognize individual... but the Gorbsky Lab is using Xenopus frog cell lines to develop robust methods for frog gene editing.... DNA at all (maybe because they have only 1,700 genes total). Hydra (Celina Juliano) This is a known stem...

...varies across targets, they tested two genes in mice. For both genes, they saw indel formation and phenotypic...certain tissues. However, packaging Streptococcus pyogenes (SpCas9) and a gRNA together (~4.2 kb) into an...al. successfully used CjCas9 with AAV to target genes in mouse muscle and eye tissue. Neisseria meningitidis...prime editors. While using AAV to deliver large transgenes might seem daunting, it’s actually a challenge...Trans-spliced Cas9 allows cleavage of HBB and CCR5 genes in human cells using compact expression cassettes...ncomms14500  Levy JM, Yeh W-H, Pendse N, Davis JR, Hennessey E, Butcher R, Koblan LW, Comander J, Liu Q, Liu...https://doi.org/10.1002/biot.201400046  Strecker J, Jones S, Koopal B, Schmid-Burgk J, Zetsche B, Gao L, Makarova...