...synthetic vectors and cultured mammalian cells. Of course, these mammalian vectors must be compatible with...discernable effect on mammalian cells. In this blog post we will discuss how mammalian plasmids differ from...supplement in mammalian cell culture to suppress bacterial growth, and is not appropriate for mammalian selection...ORI) will not allow for plasmid replication in mammalian cells, for example, and a toxin that kills bacteria...for transfected cells. Before getting into the mammalian plasmid components, it may be useful to describe...introducing genetic material (such as plasmids) into mammalian cells, a process called transfection. Transfection... and reagents vary. Plasmid transfection into mammalian cells is fairly straightforward and the resultant...

...identified. Even if some mammalian cells lines have similar morphologies, being familiar with these morphologies...Cell Culture Cloning Mammalian Cells with the Agarose Method Plasmids 101: Mammalian Vectors Resources ....org Find mammalian plasmids for your research Find viral vectors for infecting mammalian cells ...them into the incubator). It also helps to get familiar with the morphology of the cell line you’re working...)). Blood pH normally lies around 7.35-7.4 so mammalian cells are sensitive to pH. Anything between orange...24901218. PubMed Central PMCID: PMC4047077. 2. Pagliara, Patrizia, et al. "Ostreopsis cf. ovata induces...

... The Challenges of Cell Culture Plasmids 101: Mammalian Vectors Resources on Addgene.org Check Out Our...Plasmids Find Mammalian Expression Vectors ... mammalian cells... mammalian cell clones...

...can help you make an informed decision. Mammalian cells Mammalian cells are a great option for viral production...you. Table 1: Mammalian versus insect cells for viral production Consideration Mammalian cells Insect... compare two of the most widely used systems, mammalian and insect cells. Whether you’re new to in-house...production, particularly for labs that already do mammalian cell culture. HEK-293T cells are the most common... AAV, and adenoviruses can all be produced in mammalian cells. However, it is much more common to make...capacity of 8–10 kb. Time Viral production in mammalian cells is relatively quick. After transfection,... optimize for your specific application. With mammalian systems, it is standard to do only one passage...

...post we’ll provide an overview of some CRISPR mammalian expression systems, the typical applications for...those CRISPR components into your target cells.  Mammalian CRISPR expression systems Each model system has...Expression System Components of System Application Mammalian expression vector Promoter driving Cas9 expression...or stable expression of Cas9 and/or gRNA in a mammalian cell line that can be transfected at high efficiency...expression of Cas9 and/or gRNA in a wide variety of mammalian cell lines. Useful for difficult to transfect ...degraded within the cell.   Delivery methods for mammalian cell lines As mentioned above, the expression ...target cells. Delivery of these components into mammalian cell lines is quite broad and includes several...

...CRISPR article "Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9" from the Journal of...of CRISPR/Cas9 to create genomic deletions in mammalian cell lines. Below Bauer and Canver discuss the...palindromic repeats (CRISPR)/Cas9 nuclease system in mammalian cells. J.Biol.Chem. 2014 Aug 1;289(31):21312-21324... mammalian cell...

...guides mapped to a SNP would be ignored. Working with Lia Petronio and Katherine Huang from Broad’s Pattern...Javad Noorbakhsh, Isabella Boyle, Joshua Dempster, Lia Petronio, Katherine Huang, Alham Saadat, Thomas Green...

...pAquaN1 Cyan Mammalian Expresses mammalian optimized Aquamarine mCerulean N1 Cyan Mammalian Express a ...compartments pCEP4CyPet-MAMM Cyan Mammalian Expresses mammalian optimized CyPet pCyPet-His Cyan Bacterial...pCEP4YPet-MAMM Yellow Mammalian Expresses mammalian optimized YPet pYPet-His Yellow Bacterial Expresses YPet...Yellow Mammalian Express a gene of interest fused to the C-terminus of SYFP2 Clover Green Mammalian Expresses...N-terminus of monomeric Cerulean mCerulean C1 Cyan Mammalian Express a gene of interest fused to the C-terminus...C-terminus of monomeric Cerulean mTurquoise2 Cyan Mammalian Constructs to target mTurquoise2 to various subcellular...Expresses CyPet with C-terminal His tag SCFP3A Cyan Mammalian Express a gene of interest fused to the C-terminus...

...conjugated to a JaneliaFluor dye emitting at 525 nm. Image reused from https://www.janelia.org/open-science...specific targeting of microglia with AAV By Alyssa Neuhaus Genetic modification of microglial immune cells is...valuable tool for microglia research and broader therapeutic applications.  Find microglia-targeting AAV ...Voltron2 voltage sensors AAV vectors for targeting microglia The REVeRT Dual AAV Vector System Neurodegeneration... by the Schreiter Lab and the GENIE Project at Janelia, are now available! Building on their earlier Voltron...for central nervous system research; however, microglia are notoriously resistant to viral transduction...IBA1 promoter (hIBA1a, 466 bp) that drives high microglia-specific gene expression when incorporated in ...

... mind before choosing a system. Mammalian expression systems Mammalian cells are an ideal system for the...protein production and purification. These include mammalian, insect, bacterial, plant, yeast and cell free... the expression of mammalian proteins that require multiple post-translation modifications for proper ...protein function. Most DNA constructs designed for mammalian expression utilize viral promoters (SV40, CMV,...RSV) for robust expression post-transfection. Mammalian systems can express proteins both transiently ...protein yields if transfection is successful. Some mammalian systems also allow for control over when a protein...high concentrations. Despite their advantages, mammalian expressions systems do require demanding cell ...

...toward endothelial cells in rodent CNS AAV-PHP.V1 shows higher transduction in CNS endothelial cells (...exception is the X1 capsid for targeting brain endothelial cells, which is available in both AAV9 and AAV1...capsids derived from it, cross the BBB through an endothelial cell protein called LY6A (Hordeaux et al., 2019...functionally expressed on the surface of brain endothelial cells in C57BL/6J mice, but not in BALB/cJ mice...example, AAV9-X1.1 specifically transduces brain endothelial cells in mice but neurons in infant macaques ...including ENS (broad): somatic and visceral sensory ganglia, type II spiral ganglion neurons Mice, rats ...Unknown Jang et al., 2023 AAV9-X1.1 Brain endothelial cells (mice, rats); CNS neurons (infant rhesus...

...rodents, AAV9-X1.1 enables expression in brain endothelial cells via an injection into the systemic circulation... marker in the various cell types of the basal ganglia. They work via newly-characterized enhancers that...AiP13778 - pAAV-AiE0450h-minBG-iCre(R297T)-BGHpA (Alias: CN3778) AAV PHP.eB Recombinases Jonathan Ting New...AiP13779 - pAAV-AiE0779m_3xC2-minBG-iCre(R297T)-BGHpA (Alias: CN3779) AAV PHP.eB Recombinases Jonathan Ting New...AiP13781 - pAAV-AiE0452h-minBG-iCre(R297T)-BGHpA (Alias: CN3781) AAV PHP.eB Recombinases Jonathan Ting New...AiP13044 - pAAV-AiE0779m_3xC2-minBG-SYFP2-WPRE3-BGHpA (Alias: CN3044) AAV PHP.eB Controls Jonathan Ting New viral...AiP13038 - pAAV-AiE0743m_3xC2-minBG-SYFP2-WPRE3-BGHpA (Alias: CN3038) AAV PHP.eB Controls Jonathan Ting New viral...

...Similarly, the various eukaryotic cell types (mammalian, yeast, plants, etc) require unique promoters ...listing some of the most common bacterial and mammalian promoters. These lists is by no means exhaustive...considerations CMV General expression mRNA Strong mammalian expression promoter from the human cytomegalovirus... types. EF1a General expression mRNA Strong mammalian expression from human elongation factor 1 alpha...or physiology. SV40 General expression mRNA Mammalian expression promoter from the simian vacuolating...PGK1 (human or mouse) General expression mRNA Mammalian promoter from phosphoglycerate kinase gene.  Constitutive... deacetylation. Ubc General expression mRNA Mammalian promoter from the human ubiquitin C gene Constitutive...

...Vector Mammalian Lentivirus pRDA_512 (RSV-driven; lentiGuide-based) Mammalian Lentivirus... over 512) Mammalian Lentivirus pRDA_789 (CMV-driven; CROPseq) Mammalian AAV pRDA_889...AAV2) Mammalian Transposon pRDA_757 (Piggybac; constitutive expression) Mammalian Plasmid... pRDA_791 (contains only orio and ampR) Drosophilia Plasmid pRDB_051 (for stable fly cell lines...sequences for genome integration by phiC31) Drosophilia Plasmid pRDB_052 (for stable fly cell lines...integration by spontaneous insertion)   Drosophilia Plasmid pRDB_053 (for transgenic flies; gypsy...features of your vector, which are: Organism (Mammalian or drosophila) Delivery type (lentivirus, AAV...

... knockouts. Mammalian and fly applications MoClo was initially developed for non-mammalian systems, but...in bacterial and cell-free expression systems, mammalian cells, yeast, flies, and even plants! In this ...but it is not confined to that arena! The Mammalian Toolkit was engineered specifically for the types ...types of expression and integration needed in mammalian cell types, and comes with a large library of common... The EMMA Toolkit is another great option for mammalian cloning, and is geared towards cloning multi-TU...CRISPR MoClo kit termed Fragmid was developed for mammalian systems and Drosophila (McGee et al., 2023). Fragmid...The Fragmid kit described above works in both mammalian systems and Drosophila. Similarly, the Multi-Kingdom...

...community of depositors. With over 40,000 plasmids avaliable in the repository, we can't give all of them attention...molecule imaging and inducible gene expression Brilliant technologies adopt basic biological concepts and...endogenous transcription of the target gene. The brilliant SunTag system plasmids are available through Addgene...very popular. Use these plasmids to increase the brilliance of your fluorescent signal and endogenous activation...the other bits in order to achieve accurate and reliable multi-bit memory. To this end, Chris Voigt and...for light-mediated subcellular localization in mammalian cell culture and reversible control of signalling...lentiviral vector provides a powerful tool to modify mammalian cells. Multicolor CRISPR labeling chromosomal...

...AiP12237 - pAAV-AiE0452h-minBG-SYFP2-WPRE3-BGHpA (Alias: CN2237) AAV-PHPeB Control Jonathan Ting New viral...AiP12787 - pAAV-AiE0140h_3xC2-minBG-SYFP2-WPRE3-BGHpA (Alias: CN2787) AAV-PHPeB Control Jonathan Ting New viral...AiP12610 - pAAV-AiE0780m-minBG-SYFP2-WPRE3-BGHpA (Alias: CN2610) AAV-PHPeB Control Jonathan Ting New viral...pAAV-AiE0743m_3xC2-minBG-ChR2(H134R)-EYFP-WPRE3-BGHpA (Alias: CN3755) AAV-PHPeB Optogenetics Jonathan Ting New...pAAV-AiE0779m_3xC2-minBG-CoChR-EGFP-WPRE3-BGHpA (Alias: CN4033) AAV-PHPeB Optogenetics Jonathan Ting New...pAAV-AiE0452h_3xC2-minBG-CoChR-EGFP-WPRE3-BGHpA (Alias: CN4035) AAV-PHPeB Optogenetics Jonathan Ting New...3xC2-minBG-SYFP2-P2A-3XFLAG-10aa-H2B-WPRE3-BGHpA (Alias: CN4496) AAV-PHPeB Control Jonathan Ting New viral...

...biomedical model organism.  Arabidopsis thaliana Arabidopsis thaliana is one of, if not the most, recognized...introductory teaching and research environments (Idalia & Bernardo, 2017).  Escherichia coli. Image...A close relative to the mouse, both are great mammalian models due to their size, physiology, and genetic...For these reasons rats were once the most used mammalian model in medical research with an archive of information...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... of the Brown Norway rat yields insights into mammalian evolution. Nature 428, 493–521 (2004). https:/...

...revealing the unreliability of antibodies. There are three main underlying causes of this unreliability: cross-reactivity...sifting through comments and papers to find the most reliable (and cost-effective) antibody for my experiment...the antibody can no longer bind to the target). Unreliable antibodies cause research that cannot be reproduced... of all commercially available antibodies are unreliable (Baker, 2015), causing a huge waste of precious...specific (minimal binding to off-target molecules), reliable (wouldn’t perform differently over time), compatible... thousands of scientists to confidently choose reliable and consistent antibodies for specific markers...the same section and determine the antibody’s reliability through their own analysis of the background ...

...greyscale.   ARG expression in mammalian cells The first-generation mammalian ARGs (mARGs) enabled ultrasound... focused on bacterial ARGs and Tom focused on mammalian ARGs. ARG expression in bacteria The first-generation...ultrasound visualization of in situ mammalian gene expression in deep tissue. However, not unlike the first-generation...expression. We were able to express Anabaena GVs in mammalian cells without the shell stiffening protein GvpC...various cell lines from different tissues and mammalian species to produce gas vesicles. New mARG plasmids...to produce GVs in their own lab using the most reliable method, which is the transient cotransfection ... for noninvasive imaging of microorganisms in mammalian hosts. Nature 553, 86–90 (2018). Farhadi, A., ...