... 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...

...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...

...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...

...explains how gene drives work and what work he's planning on doing with CRISPR-based gene drives focusing...developing new CRISPR tools, and the potential uses of gene drives. Listen here! Podcast breakdown (Time : ...- 31:25 : Nature's defenses against gene drives. 31:26 - 33:08 : Plans for future work at the Wyss Institute... gene drives...became a member of the Addgene advisory board where he will help shape Addgene's future. Listen to this...member of the Addgene Scientific Advisory Board with a particular interest in generating Cas9 based tools...Production Specialist at Addgene. He is the audio engineer and producer for the Addgene Podcast.     Additional...

...lifecycle. Halloween genes The Halloween genes are a set of cytochrome P450 genes discovered in Drosophila...Mutations in these genes are lethal, associated with defective exoskeleton formations. These genes were appropriately...1983): 813-816. Gilbert, Lawrence I. "Halloween genes encode P450 enzymes that mediate steroid hormone...We’ve had teams dressed up as plasmids, Blugene, and the Addgene ping pong table. This year, in addition...Additional resources on the Addgene blog Read about company culture at Addgene See some of our previous ...costumes, and spooky science stories. Yes, please! At Addgene, we love finding ways to incorporate science themes...Halloween contest, we’ve also carved pumpkins (yes, Blugene made an appearance there too) and shared fascinatingly...

...mixture of XFP genes. Stochastic multicolor labeling is possible when each of the XFP genes is introduced...fluorescent protein (XFP) genes in a transgene. When multiple copies of the transgene cassette are introduced...result in a combinatorial expression of these three genes with different copy numbers, producing dozens of...modern neuroscience, plasmid or viral vector-mediated gene delivery is more common than conventional transgenic...transgenic strategies. In such vector-based gene delivery systems, the stochastic expression of XFPs is possible... Plasmid concentrations and AAV titer - The XFP genes have to be introduced at ~2 copies per cell per ...plasmids (Addgene #104102 - #104105) using in utero electroporation or with AAVs (prepared with Addgene #104110...

...varies across targets, they tested two genes in mice. For both genes, they saw indel formation and phenotypic...in vitro and germline genome editing, but in vivo gene editing approaches have been limited by problems...viral vectors (AAV) are commonly used for in vivo gene delivery due to their low immunogenicity and range...al. successfully used CjCas9 with AAV to target genes in mouse muscle and eye tissue. Neisseria meningitidis...cell is transduced by both AAVs, the full length gene and/or protein is reconstituted. One way to reconstitute...split intein spCas9-AAV toolbox that retains the gene-targeting capabilities of full-length SpCas9. This... of plasmids includes AAV-Cas9C-VPR for targeted gene activation. Split inteins have also been used to...

...knockout of a target gene and expression of a separate gene. In these mouse lines, gene knockout can be induced... type specific) control of DNA recombination and gene expression can be advanced by the coordinated use...recognizes FRT sites and initiates site directed gene recombination between those sites. FRT sites and...sequence of interest can lead to poor expression of genes within that sequence and should therefore be prevented...recombination of the loxP sites and thereby to the gene knockout. In this experiment it is possible to compare...morphology and synaptic connectivity before and after the gene knockout in the same mouse.   Scientists always...researcher exquisite control over expression of multiple genes in specific cells at specific times. Many thanks...

...CRISPR reagents and other specific gene sequences available from Addgene. Dr. McMurray hopes that by demonstrating...lab has generated mice wherein it's possible to delete the functional portion of the Nfix gene using Cre...phosphomannomutase (PMM) activity due to mutations in the PMM2 gene. Videira’s group recently reviewed and reported ...interactions and how these interactions regulation gene expression. Recent studies have shown that the conditions... Smith Syndrome are due to mutations in the NFIX gene. These diseases are characterized by overgrowth ...studying the function of the mouse Nfia, Nfib and Nfix genes in brain development since 2000 and have shown several...NFIX function. They will then mutagenize the NFIX gene and test whether variants of unknown significance...

...understanding of B. subtilis gene functions and interactions. The single-gene deletion library is derived...studying the phenotype of an essential gene, the CRISPRi Essential gene Knockdown library was constructed ...Bacillus subtilis libraries with Addgene – the B. subtilis Single Gene Deletion Library—Kanamycin and the...compatible cloning sites for easy cloning of your genes of interest. In order to improve the study of specific...cassette. However, it was unknown whether variations in gene expression from these polycistronic cassettes could... stoichiometry impacted reprogramming and global gene expression patterns. Polycistronic cassettes that... the CRISPRi Essential gene Knockdown library. Although B. subtilis has a fully sequenced genome, many...

...found in the same gene. If we only have small, directed editing abilities, any gene editing treatment ...functional copy of the entire gene, allowing a one-size-fits-all approach to gene replacement therapy for most...insertions, leaving a several kilobase-sized gap in the gene editing field. The AbuGoot lab has now developed...to work. (It was also found to be independent of gene repair.) This lowers the chance of the insert being...activity. They now had a system that could integrate genes up to 36 kb in length, with an integration rate ...system.     Fig. 2: Schematic of programmable gene insertion with PASTE. The PASTEsystem involves insertion...primary cells, and can be easily programmed for new genes. In comparison with other large insert techniques...

...EGFP can be used as positive controls for gene editing or gene activation, while the BRAF and MAP3K4 gRNAs...provide the Brunello (targeting human genes) and Brie (targeting mouse genes) pooled libraries from the labs...editing. You can also enhance expression of your gene of interest with a variety of Cas9 activators available...pooled gRNA libraries that can be used to screen for genes involved in a wide variety of cellular processes...80,000 gRNAs to target around 20,000 individual genes. We provide enough lentivirus for you to both perform...and member of the Addgene Advisory Board) says: "When you're setting up to do a genetic screen, you need... are members of the Addgene Advisory Board. Additional Resources on the Addgene Blog Get the Most from...

...Versatile Viral Tool for Gene Expression in Mammals Additional Resources on Addgene.org Find Ready-to-Use ...the cell on their way to the nucleus where their gene cargoes can be expressed. The bottom line is that...successful transduction. In another example, neuronal gene expression by an anterograde AAV was shown to drop...cell, you may not be able to detect AAV-mediated gene expression in a target cell. Practical considerations...experimental setup. Some experiments require higher gene expression and therefore higher titer than others... to such a high titer when looking for effective gene expression. How do you achieve high titers in the...therapy : the journal of the American Society of Gene Therapy. 2008;16(7):1189-1199. doi:10.1038/mt.2008.103...

...all known genes. This is a perfect example of an unbiased, reverse screen – all known genes are assayed...of knocking out the genes of interest (or the whole genome), they knock down gene expression temporarily... a biological process. Players can be proteins, genes, enzymes, drugs, or anything else that plays a role...where there is removal of known factors (such as a genes) and the loss of those factors is assessed. The ...especially useful if you are interested in studying genes whose knockout would be lethal to cells and thus...The two most common methods of manipulation are genetic knockout mutations and treatment with small molecules...selection would work is if you are trying to identify genetic conditions which convey resistance to a drug. You...

...alternative method for generating transgenic mice strains and for non-invasive in vivo gene editing. The bottom...repression, transgenic mouse line creation, and in vivo gene editing of mice, with the potential to be used for... Primary cells are often refractory to gene delivery methods, but the researchers was able to edit hiPSCs... stimulation, respectively. Primary cell type Gene targeted by guide Editing efficiency hiPSCs  EMX1...cells when using a gRNA targeting the Fto gene, while editing was not detected in cells electroporated with... used Nanoblades to disrupt the tyrosinase (Tyr) gene in mouse zygotes, which results in albinism in mice....  Nanoblades can also be used to non-invasively gene edit the liver of adult mice. When mice were IV ...