We narrowed to 932 results for: Gene

...for doing gene targeting, reporter generation, controllable gene knockouts, and inducible gene inactivation...Alec Kimmelman targets mouse metabolic genes with ~6 gRNAs per gene.  CRISPR-Act3.0, developed by Yiping... Shirley Liu's lab targets all mouse genes with 10 gRNAs per gene. The guides are optimized to maximize... to develop CRISPR-based approaches for studying gene function in these systems. As a part of this effort...Yiping Qi's lab, allows simultaneous gene activation in plants.  FnCas12a, unlike other Cas12a nucleases identified...opportunities for investigating genetic mechanisms related to morphogenesis, cell fate specification, and...Lab have compiled a new toolkit for performing genetics studies in organoids. The Organoid EasyTag System...

...multiple terminators can serve to interrupt genes (or gene clusters) in metabolic engineering. So far,...pTargetF with the ccdB gene containing BsaI restriction enzyme sites at its ends to generate pEcgRNA. The ccdB...to successfully delete genes of interest in BL21(DE3) using the newly generated pEcCas/pEcgRNA system....used MUCICAT to optimize the gene dosage of glucose dehydrogenase and generated a strain that could achieve...self-curing, an arabinose inducible lambda-red gene, and an IPTG-inducible sgRNA sequence targeting the...non-temperature-sensitive replicon, and 3) added the sacB gene for counterselection to yield pEcCas. In pTargetF... of its E. coli host, and the addition of a sacB gene renders it beneficial for plasmid clearance. It ...

...Gag, Pol, Rev, and Tat genes. Specifically for our knowledge here, the Pol gene encodes three separate...Insertional mutagenesis combined with acquired somatic mutations causes leukemogenesis following gene therapy...insertional mutagenesis. This integration could lead to an undesirable interruption of a critical gene. Because...Insertional mutagenesis is especially concerning in clinical settings, as evidenced by retroviral gene therapy...transcriptase), and IN (integrase). These viral packaging genes are required for the production of lentiviral particles...portion does not contain any of the viral packaging genes, the natural cycle of the lentiviral vector ends...transcriptionally active, leading to transient expression of gene products (Wu et al., 2004). Over time, the expression...

...2020. . “Advances in CRISPR/Cas-based Gene Therapy in Human Genetic Diseases.” Theranostics. https://doi.org...treat AD using CRISPR/Cas9. They are targeting the genes responsible for both early onset and late onset ...Sciences found that mice with a dominant mutation in a gene that causes cataracts could be rescued by coinjection...dominant cataract disorder caused by a single copy of a gene known as Crygc. When all was said and done, the ...able to demonstrate functional correction of the gene in clonally expanded organoids. “CRISPR in adult...fibrosis may not be the most likely candidate for gene therapy, however, because the disease affects multiple...proof-of-principle for use of CRISPR to correct single-gene conditions. Remaining hurdles in CRISPR biology...

...either side of the cargo (ex: fluorescent protein gene) on the donor vector. Therefore, targeting requires...developed one of these systems further for simultaneous gene activation and editing. Read the paper in Nucleic... with an NGG PAM motif, averaging 19 targets per gene. The lab has previously used the library with SpdCas9...perform pooled CRISPRi screens to identify essential genes and phage host factors, and identify sequence-specific...Knockout Library is a lentiviral library targeting 26 genes chosen from an analysis of human metastasis cohorts...metastasis CRISPR screen. The Chen Mouse Tumor Suppressor Gene CRISPR Knockout Library is an AAV CRISPR knockout...knockout library targeting mouse tumor suppressor genes (TSG). Find the Mouse Metastasis CRISPR-Cpf1 Double...

...reporter gene (e.g., mCherry) to easily detect opsin-expressing cells. The resulting fusion gene is inserted...downside in optogenetics (Belzung et al., 2014). This could be overcome by fusing opsin genes to cell-type...Krook-Magnuson, 2018). Optogenetic FLEx vectors contain a strong promoter upstream of an opsin gene (e.g., channelrhodopsin...2014). Microbial rhodopsins are encoded by opsin genes, and include bacteriorhodopsins, halorhodopsins,...light-sensitive, so the selective expression of opsin genes in targeted cells makes it possible to control neuronal...viruses (rAAVs) have become increasingly popular for gene delivery in the central nervous system due to their...those usually drive weak expression of downstream genes. Strong cell-specific promoters, on the other side...

... on a break from expressing some genes of interest).   Blugene is so excited they can’t stop dancing...GIFs, feel free to let us know on Twitter @addgene! Addgene is always looking for new ways to help you...We’re making some moves over here at Addgene...and they’re looping? What’s the deal with looping...coworkers, labmates, and friends as well!   Addgene is sharing so much more than plasmids these days...GIPHY   You can see for yourself at giphy.com/addgene. But that’s not all! You’ll be able to use these...Twitter, Instagram, TikTok and more. Just type “addgene” into the GIF search bar to find that perfect response....   This could be you after watching Addgene’s protocol videos. via GIPHY   If you’re also...

...simple example with the goal is to generate mice that have two modified genes in their genome, and assuming...your targeting vector or gene of interest. 2. Breed your parental mice to generate F1 Once you’ve performed...other will generate approximately 1 in 16 mice that harbor homozygous alleles at both of your genes of interest...your mice. Because their generation involves the integration of a novel gene randomly into their genome...your parental mice were all homozygous for your genes of interest, your F1 mice will be carriers at those... all of your desired genes. Remember Punnett squares from high school biology? They will come in handy...expresses Cre with a strain that expresses a floxed gene of interest. Cre may be expressed globally (in every...

...antibiotic resistance genes, as well as non-selectable DNA fragments, such as gene replacements and tags...repressor gene, CI857, is used.  This mutant repressor prevents expression of the recombination genes at low...the red genes needed for recombineering, but also leads to expression of the int and xis genes which are...Substrate DNA design and generation Expression of lambda red recombination genes. Electroporation of substrate... a different genetic background via P1 transduction. Plasmid Expressing lambda red genes from a plasmid...recombinant clones are generated.  A simple way to do this is to express lambda red genes from a plasmid with...using Lambda Red recombineering system to replace a gene of interest with an antibiotic resistance cassette...

...number of positive droplets for the gene of interest and reference gene, copy number can be determined through...your experiment?  Complexity of the gene of interest Some gene sequences are harder to amplify and quantify...is a well known fact if a multicopy variant of a gene exists, there must be assays that researchers can... ubiquitously expressed, one-copy ‘housekeeping’ gene), and quantified relative to a 'normal' sample with...to use a reference sample with two copies of the gene (Ma, 2014.) As with all qPCR assays, samples, standard...between 0.80-1.20) would mean you have one copy of the gene; an RQ value of 2 means you have two copies, and... assays, allowing the reference sequence and the gene of interest to be amplified in the same reaction...

...disease in which disruption of the smn1 gene results in the degeneration of spinal cord motor neurons. This...Schiavi will use Addgene plasmids to characterize smn1 interactions with other genes. He will use fluorescent...to identify candidate genes as drivers of peritoneal mesothelioma tumorigenesis. They plan to use CRISPR...assess their tumorigenesis. These experiments will serve to validate these candidate genes and allow Nabavi...19,000 live births (1).  While disruption of the smn1 gene is known to cause spinal muscular atrophy, the molecular...fluorescent proteins to monitor the localization of these genes, CRISPR to disrupt their function, and C. elegans... Di Schiavi will be able to better choose which genes and pathways to target in future small molecule ...

...and can deliver large sequences and even entire genes. Figure 1: Mechanism of CAST systems. A Cas...evolution using bacteriophages. Researchers moved a gene required for phage propagation to a plasmid that... CAST integration of the promoter could turn the gene on. Once phage success was linked to CAST efficiency...new horizons of evoCAST In the emerging world of gene therapy, highly tailored treatments to specific ...In contrast, a system that can deliver a healthy gene as a single intervention for a variety of loss-of-function...evoCAST is an important step toward that kind of gene treatment. The researchers also compared evoCAST... (Prime-Editing-Assisted Site-Specific Integrase Gene Editing), which combines prime editing with a site-specific...

...interspaced palindromic repeats) Cas (CRISPR associated genes) system. This popular genome engineering tool offers... be incorporated into the bacterial genome. Cas genes, which code for polymerases, nucleases, and helicases...adaptive immune system that utilized various cas genes not only to store a record of invading phages but...conserved CRISPR subtypes comprised of more than 40 cas gene families have been characterized. To date, 83% of...Rising to the challenge - Evolution of anti-CRISPR genes in phages The CRISPR/Cas adaptive immune system ...nucleases from re-identifying it (3). Surprisingly, few genes have been identified that neutralize CRISPR/Cas....Earlier this year, expression of 1 of 5 anti-CRISPR genes were found to inactivate the Type I-F CRISPR/Cas...

...variants in a gene of poorly classified function and a well established cellular signaling gene, thus providing...above to demonstrate how gene expression profiling, systematic tumorigenesis assays, drug epistasis assays... be used as generalizable strategies to map the function of rare cancer variants and genes: High-throughput...bottleneck is that most recurrently mutated cancer genes are rare and most of the individual variants found...well-annotated cellular pathways). It wasn’t clear whether gene-agnostic bioassays could accurately test for molecular...numbers of cancer variants across a large number of genes as systematically as possible. In developing these...assays were designed to complement the one assay per gene approach that has been the historical standard. ...

...them together. Gene expression The reason for most viral experiments is to deliver a genetic cargo, typically...primary plasmid with the delivered cargo (often a gene) is called the transfer plasmid while the other ...component as well. On both sides of this promoter-gene combo are two long terminal repeats (LTRs). These...infiltrate the host cell and deposit viral cargo. Gene expression While AAV often does not integrate into...(more on this below), it’s still used to deliver gene expression constructs. Sandwiched between two inverted... AAV also requires some back up from adenovirus genes – E4, E2a, and VA – which are supplied on a helper...viral replication. More information on these viral genes is available in the adenovirus section below!   ...

...plasmid in the host bacterium that expresses gene III. Gene III is under the control of a T7 RNA polymerase... the expression of gene III to create infectious particles. A plasmid-based genetic circuit in the bacteria...activity. A plasmid-based genetic circuit in the bacteria links the expression of gene III to base editor activity... 1: David Liu gives a talk at the Cell Symposia: Gene- and Cell-based Therapies: CRISPR, Stem Cells, and...Cas domains beyond SpCas9. In order to expand the gene-editing applications and targeting scope of ABEs...bacteria links the expression of gene III to base editor activity and editing speed and consists of the... active variant of the base editor, a functional gene III product is produced and can infect bacterial...

...the ccdB gene is replaced by the gene of interest. As a result, cells that retain the ccdB gene die off...tool. The plasmid pKSCTMF contains the mazF gene and a gene for Zeocin resistance (Figure 2). This MazF-ZeoR...antisense RNA regulates hok gene expression indirectly through the overlapping mok gene." Journal of molecular...associated genes. These systems better ensure that plasmids are passed down through generations (Figure ...application of CcdB is GatewayTM cloning, where the ccdB gene serves as a counterselectable marker in the destination...death. The team used this cassette to knock out genes, knock in a GFP cassette, and introduced site-directed...toxin-antitoxin system to construct a stable synthetic gene circuit that can be used over prolonged periods....

...Express a gene of interest fused to the C-terminus of SCFP3A Amber N1 Yellow Mammalian Express a gene of interest...Express a gene of interest fused to the C-terminus of Amber mVenus N1 Yellow Mammalian Express a gene of interest...your choice of fluorescent fusion protein with your gene of interest: Plasmid Color Expression Description...Aquamarine mCerulean N1 Cyan Mammalian Express a gene of interest fused to the N-terminus of monomeric...Cerulean mCerulean C1 Cyan Mammalian Express a gene of interest fused to the C-terminus of monomeric...monomeric Venus mVenus C1 Yellow Mammalian Express a gene of interest fused to the C-terminus of monomeric...C-terminal His tag SYFP2 Yellow Mammalian Express a gene of interest fused to the C-terminus of SYFP2 Clover...

...activators and repressors to target genes. Repressing target genes using dCas9-based repressors Early ...of the gene of interest. dCas9-based repressors are useful when knocking out a particular gene isn’t feasible...repressing target genes in a variety of species and cell types can be found on Addgene’s website. Activating...activating target genes in a variety of species and cell types can be found on Addgene’s website. ...Designing or selecting a sgRNA Addgene may carry a validated sgRNA for your target gene.  Validated gRNAs are ... activate, repress, and even image your favorite gene. With so many Cas9 reagents available, it can be...introducing Cas9-mediated frameshift mutations in the gene of interest. The genomic target for Cas9 and the...

... this gene. Should you cut it at the beginning of the coding sequence to delete the entire gene? Perhaps... in vitro, and using it in mouse embryos to generate gene modified mouse lines. Hopefully this post will...they have been generated (more on that later). Find Validated gRNAs to Target Mouse Genes To begin the ...chances of gene modification in vivo. Find Plasmids for Testing Your gRNAs in vitro at Addgene CRISPR/Cas9...decide to do a gene modification experiment in mice to the day you have your F0 generation (barring life...Haoyi, et al. "One-step generation of mice carrying mutations in multiple genes by CRISPR/Cas-mediated ...blogger Samantha Young. The use of CRISPR/Cas9 for gene editing has expanded since its adaptation for use...

...knockout, activate or repress target genes in plants using the same general experimental design principles ...Resistance by CRISPR/Cas9-Targeted Mutagenesis of the ERF Transcription Factor Gene OsERF922. PLoS ONE 11:e0154027...for targeted modification of known or suspected genes that regulate a desired phenotype. In fact, CRISPR... can also be used to activate or repress target genes in plant cells, respectively. Cas9 expression is...regulate expression of longer RNAs (such as mRNAs for gene expression). Examples of commonly used RNA pol II..., activation and repression of target genes in plants and many of the aforementioned empty gRNA backbones...Agrobacterium tumefaciens as a vehicle to deliver your gene of interest into a target cell line or organism ...

..."Future of rAAV gene therapy: Platform for RNAi, Gene Editing and Beyond." Human Gene Therapy ja (2017...required for the onset of gene expression. However, it has yet to be determined what gene expression looks like...Additional Resources on the Addgene Blog AAV: A Versatile Viral Tool for Gene Expression in Mammals A Match...transport desirable or not desirable? If diffuse gene expression is tolerated, then axonal transport is...Otherwise, using a specific system for controlling gene expression such as Cre-lox, may be sufficient to...tool to use. However, we are starting to see that gene expression really does not take much time at all... normally require second strand synthesis before gene expression begins. ScAAV avoid this rate limiting...

... to make my first CRISPR/Cas9 gene edit. I wanted to inactivate the gene BRAF (a kinase implicated in ...Targeting different regions of the gene is also useful in case the gene has a weird mutation that you didn...each step of performing and analyzing a gene edit using Addgene’s lentiviral CRISPR tools. Schematic...Schematic overview of CRISPR/Cas9 gene editing process. See below for details. The entire process from start...what level of expression was the most efficient for gene editing. A virus encoding GFP plus a gRNA targeting...GFP gRNA, will edit out the virally-delivered GFP gene. So, cells with no Cas9 should appear green and ...multiple gRNAs to target different regions of your gene. I used existing BRAF gRNA viruses. There are a ...

...especially when fused to effector proteins for gene activation or gene editing. The more recently identified Cas12f...SEND for gene transfer with pseudotyped viral particles, expanding options for CRISPR or other gene delivery...editing systems improve gene editing performance at therapeutically relevant gene targets and expand the...version (named CasMINI) had the highest activity for gene activation  and base editing, whereas a different..., and fusogen in a modular system for delivering gene editing tools into cells. Image adapted from Segel...editing to correct mutations in a variety of target genes and cell types. Their Nature Biotechnology paper...tyrosinemia, where a pathogenic insertion within the FAH gene was precisely corrected, restoring expression in...

... transcription of a target gene, it therefore determines the timing of gene expression and largely defines...synthesis and allowing reporter gene transcription to take place. Addgene also has popular light-regulated...the promoter and ceases repression of the target gene. A simplified lac inducible system is found in many...shock-inducible Hsp70 or Hsp90-derived promoters, in which a gene of choice is only expressed following exposure to...inducible promoters Light is another way to activate gene expression, and two-component systems used in synthetic...promoter pR, preventing expression of a reporter gene. When light is present, YFI is inactive, preventing...experimental system. Leakiness If you’re expressing a gene that may be toxic, it’s important that your inducible...

...the plasmid containing the lethal gene. An example is the ccdB gene system commonly used in Gateway® Cloning...are rescued by genes supplied on the transformed plasmid. Such plasmids may contain genes that allow for...auxotrophic cell lines), or genes which rescue a lethal phenotype. Resources on the Addgene Blog Read Our Post...well-characterized bacterial lac operon contains a gene called lacZ that encodes for the enzyme β-galactosidase...discovered that deleting a section from the lacZ gene (a mutation called lacZΔM15) creates a non-functional...other methods. Positive selection vectors encode a gene which, when expressed, is lethal to the cell. Cloning...fragments are inserted into an MCS in the center of this gene, disrupting the lethality. This is similar to α-...

...number of genes involved in cardiovascular and metabolic diseases. We are knocking out genes that have...and find several mutations in the same gene or in multiple genes, all of which seem to contribute to a ...versions of 15 different disease-associated genes. Addgene spoke to Musunuru about how the new kit works, ...scenarios, you can either knock out a gene or knock in a mutation. Addgene: How do you use this kit to target... paper, for example, we generated hepatocytes to examine the effect of a gene on cholesterol metabolism...They do that by studying patients to uncover new gene variants associated with conditions of interest,...be advantageous if you are trying to knock out a gene. One can also exploit homology-directed repair by...

...used in the lab, the transposase gene is provided in trans so that a gene of interest can be inserted between... 2021).  Gene transfer Transposons are an alternative to viral vector delivery of transgenes, like for...response and are also easier and cheaper to generate. Gene disruption due to integration can occur for...several different applications (mutagenesis screens, transgenic animals, gene transfer). The discovery of ...bacteria, transposons can move antibiotic resistance genes between plasmids or from plasmids to the bacteria... genome. In humans, if a transposon jumps into a gene, it could be mutagenic and result in human disease... and mechanism of replication. They contain two genes: gag and pol. The pol polyprotein encodes the reverse...

...to investigate the roles of specific genes or to knock down genes with potentially harmful mutations in...target gene but have a lot of off-target effects. However, this is a common issue with many gene silencing...this could be applied to. When it comes to altering gene expression, CRISPR technologies have gradually taken...) is, as the name might suggest, an RNA-mediated gene silencing mechanism. RNAi was a landmark discovery...focusing on shRNA, one of the most popular choices for gene silencing. RNA interference RNAi is a useful tool... in therapeutic settings. Silencing of a target gene is achieved through the delivery of a double-stranded... these tools. Next time you need to knock down a gene, maybe you’ll consider using RNAi! References and...

...specific genes are signified with a minus sign (-) as is typically standard; just having the gene listed... of the more common genetic changes found in E. coli strains. Table 1: Common gene mutations found in ...carefully selected the optimal promoter for your gene of interest, painstakingly cloned into the perfect...backbone, made sure to add the right tags to your gene, and may have even put a fluorescent protein downstream...in E.coli strains Gene(s) Description Functional Consequence dam DNA adenine methylase mutation (GATC...methylation sensitive. dnaJ Mutation in a chaperonin gene Increases the stability of certain expressed proteins...encodes proteins needed for bacterial conjugation. Genes listed on F´ are wild-type unless indicated otherwise...

...aberrant activation of nearby genes. These safer vectors have become standard in gene therapy. Researchers have...condensed to create the 2nd-generation lentiviral system (Figure 2). The HIV genes that do remain are very...into the genome, they could promote oncogenesis by altering local gene expression. Self-inactivating (SIN... LM, Rebello T. J Gene Med. 2013 Feb;15(2):78-82. PubMed PMID: 23322669. 6. Genetic treatment of a molecular... Jr. Gene Ther. 2010 Feb;17(2):150-7. PubMed PMID: 19847206. Additional Resources on the Addgene Blog ...applications, and they’ve also had success in clinical gene therapy applications. Read on to learn more about...researchers are interested in developing these vectors for gene therapy. In 2003, VIRxSYS began the first clinical...

...MSCV (Murine Stem Cell Virus) sequences. Packaging genes (viral Gag-Pol): Gag is a structural precursor protein... protein, and Pol is a polymerase. Envelope gene (may be pseudotyped to alter infectivity): The use of...both lentiviruses and γ-retroviruses use the same gene products for packaging (Gag, Pol, and Env), the ...and/or Env, eliminating the need to deliver these genes in trans by specific plasmids. This method, therefore...standard γ-retroviruses use the Gag, Pol, and Env genes for packaging; however, they are different viruses...replication incompetent? Yes, as with lentiviruses, the genes required for γ-retroviral packaging and transduction...U3 region of the 3’ LTR and are primarily used in gene therapy research to reduce promoter interference...

...multiple gRNAs targeting a single gene can increase the likelihood of gene knockout, or multiple gRNAs may...genome editing has been used to study the effect of gene knockout in vivo and in vitro, as well as to insert...observe a phenotype, the modification of multiple genes is often necessary. Guide RNAs (gRNAs) are commonly... al constructed polycistronic glycine tRNA-gRNA genes (PTGs) in the forms tRNA-gRNA and tRNA-gRNA-tRNA...constructed PTGs containing 8 gRNAs targeting 4 MAPK genes, again using rice protoplasts. Each pair of gRNAs...may be used to target genes with overlapping activity. Multiple gRNAs can also facilitate deletions of...viruses have illustrated the power of polycistronic gene organization, and its translation to CRISPR/Cas9...

... or “LSL” cassette) upstream of a gene of interest will prevent gene expression in the absence of Cre....Cre-dependent gene knockout – conversely, putting the loxP sites on either side of a gene (called “floxing... loxP”), will permit gene expression until Cre is present, at which time the gene will be disrupted or...most common uses are listed below. Cre-dependent gene expression – placing a series of transcriptional... presence of Cre, the stop codon is excised, and gene expression proceeds. One popular lox-stop-lox plasmid...switch regions evidenced through Cre–loxP–mediated gene targeting. Gu, H., Zou, Y.R., and Rajewsky, K. 1993...reciprocal crossover event. This event can have three general outcomes based on the location and orientation ...

... (NHEJ) pathway disrupt target gene sequences (thus inactivating genes), while breaks repaired by homology...error-prone repair of the gene would be disastrous (i.e. if breaking the gene is worse than leaving the...(in our hands) this technique supports gene disruption or gene replacement frequencies that are an order...repair (HDR) pathways can modify the sequence of a gene (thus altering its function). HDR is crucial for...consistently supported higher frequencies (up to 60%!) of gene editing than single or double stranded donor DNA...editing, but it could be very useful to tackle genetic diseases in which sequence replacement holds some...PubMed Central PMCID: PMC4032847. Resources on the Addgene Blog Brush Up on Your HDR Basics Learn How to ...

... delete/overexpress endogenous genes or to introduce heterologous genes, but the trend of research has...tinkering with regulatory networks and multiplex gene control. However, these approaches are often met...mimic natural transcriptional programs on multiple genes. Modifying the CRISPRi system for multidirectional...programs to differentially alter the levels of five genes that encode biosynthetic enzymes for violacein, ...recruit various transcriptional regulators to alter gene expression at the targeted site. Multiple scaffold...simultaneous, multidirectional and tunable control of gene expression. This strategy can be applied to effectively...RNA-guided platform for sequence-specific control of gene expression. Cell, 152(5), 1173-1183. (2013). PubMed...

...profiles (Dwight, 2011).  Reference Genes  Choosing a set of reference genes deserves more attention than it...reference genes, plural. Your data will be much improved if you move to using multiple reference genes whenever...reference gene being a common choice - the frequently used GAPDH has noted issues as a reference gene (Higgins... Validation of Common Housekeeping Genes as Reference for qPCR Gene Expression Analysis During iPS Reprogramming... it usually receives. Different reference genes have shown to have as much as sixfold variation in expression...), so you’ll want to ensure that your reference genes are suitable for your cell lines and/or tissue types..., and it's possible to multiplex your reference genes to save on plate space. For many cell lines and ...

...easily modified to tag hundreds of genes, generating powerful genetic screening libraries. Additional protein... to screen for the genetically modified strains that contain your newly tagged gene. Most insertions can... to generate high-throughput targeting vectors. These vectors can be used to tag endogenous genes and ...-Throughput CRISPR/Cas9 Gene Modification in Caenorhabditis elegans.” Genetics. 202(4) (2016):1277-88....protein fusions is to replace the genomic copy of your gene of interest with a fluorescent protein fusion. CRISPR...allowed scientists to tag and light up endogenous genes of interest to better understand normal protein ...easily create C-terminal tag fusions of endogenous genes in human cells. To use CRISPaint, users need 3 vectors...

...that of adjacent cellular genes. Q7: Which genes are deleted or modified to result in replication deficiency...including the gene of interest, and the plasmid coding for an envelope. Three generations of lentiviral...minimal number of HIV genes required for virus production (3 or 4). Third-generation vectors contain gag...gag, pol, and rev. Second-generation contains those three genes plus tat. The envelope plasmid provides...are useful and efficient tools to introduce your gene of interest into cells. Unlike gamma-retroviruses...lentiviruses and gamma-retroviruses use the same genes for packaging (that's gag, pol, and env), the isoforms...infect dividing cells. When choosing the appropriate gene delivery system, the specific target cell or tissue...

...forward engineer a system of 10 genes. You can now forward engineer a 10-gene system to do something. We know...collection of genetic parts for precisely forward-engineering the central dogma across many genes simultaneously..., off-the-shelf genetic elements that allow you to begin to imagine designing 10-gene systems with quantitative...used over and over again in combination with novel genes to get very precise expression levels. The BIOFAB...otherwise invisible molecules into regulation of gene expression including growth or fluorescence. There...our testing of these parts that the chance your genes will be expressed at the level you want is 93 percent... of biological systems today who are building 10-gene systems from scratch. (They may be building genomes...

...physiological gene expression regimes that cannot be easily achieved with classical gene overexpression...contain changes in expression of TRIM21-associated genes, indicating that TRIM21 overexpression does not ...variety of protein modules to repress or activate gene expression, sometimes incorporating chemically inducible... 28916764. Plasmids for independent control of gene expression in yeast Article contributed by Tyler...that can independently control expression of your genes of interest. These transcription factors consist... appropriate promoter sequence and activation of gene expression. In addition to plasmids encoding these...and ZIF268. To demonstrate independent control of gene expression, Aranda-Díaz et al transformed yeast ...

...their test genes, the researchers at the BIOFAB showed that expression of many different genes was predictable...translational repressor to down-regulate a gene of interest. You can generate PUF proteins with a wide variety ...restriction enzymes and standardized primers to clone your gene of interest into both sequencing (pInitial) and ...abundance (meaning they are unlikely to be found in your gene of interest) and prevent incorporation of extensive...kit contain a variety of promoters to facilitate gene expression in bacteria, yeast, mammalian cells, ...hope this kit will allow you to quickly study your gene of interest in a variety of model systems, thereby...peptides) downstream of which you can clone your gene of interest. Translation of the standardized leader...

...viral (AAV) vectors are the most frequently used gene-transfer tools in the study of the brain and spinal...organs to the CNS, are also an important target for gene delivery, especially for the study of pain. While...all possible combinations into a region of the cap gene. In this special CREATE cap plasmid, loxP sites ...primers to determine which version(s) of the capsid gene successfully infected the cell type of interest....translational applications (mechanistic studies and gene therapy treatments for neurological diseases). The...capsids can serve as powerful cell-type-restricted gene vectors for neurons, no transgenic mouse required...et al. Engineered AAVs for efficient noninvasive gene delivery to the central and peripheral nervous systems...

... what part of a gene enhancers like to interact with, and how many promoters or genes each enhancer is...plot line in gene regulation. When chromatin conformation data is compared with changes in gene expression...dynamic role in gene regulation The high resolution and multiplexed chromatin ‘photos’ generated with CAPTURE...changes in gene expression and determine who’s a key driver of these changes. By generating a more vivid...updated Jul 2, 2020. Promoters may be the star of gene regulation, but enhancers and chromatin looping ... bound by transcription factors, can increase a gene’s transcription. Sometimes enhancers are located ...located thousands of base pairs away from the gene they regulate, but are brought in proximity by the looping...

... one bacterium sharing genes with another, an accidental transmission of genes from one bacterium to another...‘horizontal gene transfer’. Resistance can also arise from random mutations in the genetic sequence that...Additional Resources on the Addgene Blog Learn How Antibiotic Resistance Genes Are Used in the Lab Probe...consuming to treat. Learn How Antibiotic Resistance Genes Are Used in the Lab How does antibiotic resistance...bacterial cells), or a bacterium picking up resistance genes from it’s environment (from DNA left behind after...develop, either through random mutation or horizontal gene transfer, means to defend themselves from these ...while the few that happen to possess resistance genes survive and multiply. Factors contributing to the...

...resistance gene, allowing resistant bacteria to multiply, spreading their resistance genes throughout ...them to multiply and propagate their resistance genes. Our society is hoping to combat antibiotic resistance...bacterial species, or even an antibiotic resistance gene. Importantly, native, non-pathogenic bacteria remain.... targeted beta-lactam and quinolone resistance genes in E. coli using plasmid and phagemid delivery systems...population to the antibiotic, despite this resistance gene being encoded by a high-copy plasmid. In the case...CRISPR machinery targeting 1) the given resistance gene and 2) a second lytic phage. Bacteria that receive...work targeted two distinct beta-lactam resistance genes, again with very high specificity. This double phage...

... containing structural genes and a transfer plasmid carrying the transfer gene. By dividing the viral ...Talking 'bout my (lentiviral) generation The first step in delivering genes via lentivirus is deciding what... While lentiviral vectors are popular gene delivery tools, producing lentivirus, can pose certain challenges...split the viral genome by removing the viral Rev gene from the packaging plasmid and incorporating it ... the structural elements, envelope, and transfer gene are transfected into a packaging cell line, usually...) while a single 72h harvest is best for larger genes such as Cas9. See figure 1 for an example of how...sufficient titer; this is often the case with large genes. This is due to inefficient packaging of large inserts...

...screens using CRISPR technology, gRNAs to knockout genes – are essential screening components, the model ...Sometimes, you might have a few positive control genes on which to test the validity of your biological... a new assay or both, and you have no idea what genes are involved. Or you don’t know how well your assay...definition, the kinome is a well-annotated collection of genes that can provide a useful starting point for developing...screens can be performed with either 4 or 8 gRNAs per gene. Further, the pools are available in two vector ...individual plasmids will also be useful for imaging or gene expression assays necessitating the screening of...blogger, member of the Addgene Advisory Board, and Associate Director of the Genetic Perturbation Platform...

...was previously a large barrier for gene therapy. In fact, AAV gene therapies have already been approved...Transient Gene Expression High Immunogenecity AAV Remains Episomal (Little Risk of Mutagenesis) Very...of Insertional Mutagenesis) High Gene Expression Infects Non-Dividing Cells Large Packaging Capacity (...Resources on the Addgene Blog Learn More about Lentivirus Listen to Connie Cepko Discuss Gene Therapy with...Type of Virus Pros Cons Retrovirus Stable Gene Expression Broad Tropism Only Infects Dividing...Immunogenicity Infects Non-Dividing Cells Transient Gene Expression Low Tropism for Mouse Cells and Hematopoietic... lentiviral vectors through pseudotyping.” Curr. Gene. Ther. 5 (2005) 387-398. PubMed PMID: 16101513. ...

...of homologous recombination to facilitate simple gene replacement/mutation. Yeast expression plasmids used...host mutations by including a copy of a functional gene which complements the host’s auxotrophy. When grown...have incorporated the plasmid carrying the required gene.  The table below lists some of the most commonly...growth-based positive selection for the loss of the marker gene) For use in: Can this be used in other yeast species...yeast phenotypes. This marker should not be used in gene disruption experiments. MET15 L-methionine and...FOA in fission yeast induces mutation in the ura5+ gene in addition to ura4+. leu1+ L-leucine   S. pombe...issues. One method to reduce the amount of marker gene expression is to use a partially defective promoter...