... Plasmid Collections Mammalian RNAi Mammalian shRNA Tools for RNAi Constitutive Lentiviral...Plasmids for mammalian RNA interference (RNAi) experiments at Addgene; browse by constitutive lentiviral...RNA-mediated gene silencing mechanism. As a tool in mammalian cell systems, silencing is achieved through the...system to functionally silence genes in primary mammalian cells, stem cells and transgenic mice by RNA interference...

...synthetic biology plasmids for use in mammalian systems. Mammalian Plasmids Search the table by keyword...plasmids for use in mammalian systems. Plasmid... Plasmid Collections Synthetic Biology Mammalian Synthetic Biology...Biology: Mammalian SynBio Resources SynBio Home Browse All SynBio Assembly Standards SynBio Blog Genome Engineering...

... CRISPR Mammalian Expression CRISPR Plasmids: Mammalian Expression The... that function in mammalian cells can be used to attentuate RNA levels. In mammalian systems, Cas13a does...or cloning method. To isolate mammalian vectors, simply type "Mammalian" into the search bar. When using...designed for use in mammalian systems. CRISPR...CRISPR plasmids have been designed for use in mammalian expression systems. Cut Fully functional CRISPR...Libraries gRNAs Empty gRNA Vectors gRNAs By Species Mammalian Bacteria Drosophila Plant C. elegans Yeast Zebrafish... C->T (or G->A on the opposite strand) within a small editing window specified by the gRNA. Adenine base...

...Genomic Deletions in Mammalian Cell Lines CRISPR: Protocol for Genomic Deletions in Mammalian Cell Lines Other...by a small indel at the sgRNA target site. Design additional primers to analyze for scarring (small indels...Experiments (JoVE) about genomic deletions in mammalian cell lines using CRISPR/Cas9 CRISPR...of CRISPR/Cas9 to create genomic deletions in mammalian cell lines. Along with the video, you can find...Article Citation Generation of Genomic Deletions in Mammalian Cell Lines via CRISPR/Cas9. Bauer DE, Canver MC...potential advantages as compared to single-site small indels given the efficiency of biallelic modification...loss-of-function studies of genes and genetic elements in mammalian cell lines. Protocol For referenced Tables and...

...compartment (chloroplastic, cytoplasmic, glyoxysomal, mitochondrial, peroxisomal) Mutation (specific mutation, "...Browse a diverse set of wild-type malate dehydrogenase genes and watermelon mutant clones for your course-based... Depositor Collections MDH CUREs Community Malate Dehydrogenase CUREs Community (MCC) Collection ...Experiences (CUREs) (Link opens in a new window) The Malate Dehydrogenase CUREs Community (MCC) (Link opens...Experiences (CUREs) centered on a single enzyme, malate dehydrogenase (MDH). The MCC is a national group...Visualization to Develop Student Hypotheses in a Malate Dehydrogenase Oriented CURE. Callahan KP et al....learning and attitudinal outcomes: A study of the Malate dehydrogenase CUREs Community (MCC). DeChenne-Peters...

...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...Although plasmids do not naturally exist in mammals, scientists can still reap the benefits of plasmid-based...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...

...yellow) - 8.0 (red)). Blood pH normally lies around 7.35-7.4 so mammalian cells are sensitive to pH. Anything...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 ...flowing in. The vent for the in-flowing air is normally visible and you should always ensure nothing is...cross-contamination can be identified. Even if some mammalian cells lines have similar morphologies, being familiar...

...because of a small, avoidable mistake. Or worse, when you're trapped in a pattern of small errors that ... when you’re in the lab and you're sweating the small stuff? Identifying the Issue You can’t fix an experiment...your prep an investment in accuracy.  Bench setup Small adjustments in the physical organization of your...reduce errors as you work, and the possibilities for small, helpful changes are endless.    Fig. 2... Just like any other experimental approach, use small, incremental changes to understand what works and...

...each clone in case of contamination. We do not normally use antibiotics. Lastly, and perhaps obviously... 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...multiple different cell types, including plants, mammals, and insects. In this post, we will compare two...two of the most widely used systems, mammalian and insect cells. Whether you’re new to in-house virus production...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,...

...Born to a family in Malawi that pushed science education, Kingdom Kwapata grew up to be one of the best...among 100,000 eligible to attend the University of Malawi where he focused on agricultural engineering through... Kingdom on opportunities to go into science in Malawi 4:36-5:46 : How Kingdom ended up working in the...University 9:36-10:29 : Kingdom's transition back to Malawi and Lilongwe University of Agriculture and Natural...difficulties surrouding and need for biotech research in Malawi 11:41-14:53 : Kingdom's current projects producing...projects - HIV inhibitors, biofuels, challenges in Malawi, and more 23:45-28:07 : How Seeding Labs is helping... malawi...

... design and optimize. Today we’ll cover the normalization and loading controls needed for relative quantification...In relative quantification, the POI bands are normalized to the loading control: the intensity of each...resulting fraction is reported. Figure 2: Normalization with loading controls. Under each lane, the ...the loading controls are equally strong, so the normalized value simply reports which POI band is stronger...is weaker in condition 1 than condition 2, the normalized values for the two conditions are equal. Created...Created with BioRender.com. This kind of normalization is a nearly ubiquitous tool, but proceed with...your western blot, and depending too much on normalization can amplify other errors in experimental design...

... also confines channelrhodopsin activation to a small volume of brain tissue (~1 mm3). Current channelrhodopsins... To make channelrhodopsin-based optogenetics minimally-invasive, Frances Arnold’s lab and Viviana Gradinaru...channelrhodopsins paired with a systemic AAV allow for minimally-invasive optogenetics While ChRgers performed ...in mice to see if they were suited for in vivo minimally invasive optogenetics. Both experiments delivered...paired with PHP.eB’s systemic delivery enables minimally-invasive gene delivery and optogenetic excitation...learning-guided channelrhodopsin engineering enables minimally invasive optogenetics. Nat Methods 16:1176–1184...

...replicated many of the properties once exclusive to small-molecules: red-shifted spectra, ion sensitivity,...improve the photon budget. Of course, “reverting” to small molecule dyes seems daunting—no one wants to spend...that reacts specifically and irreversibly with a small substrate ligand motif attached to a fluorophore...These modified antibody fragments bind and enhance small molecule fluorogens (Figure 1e). Stains – These..., no genetically encoded tag is required as the small-molecule binding motif targets endogenous protein...the system. A particularly desirable property for small-molecule labeling strategies is fluorogenicity, ...flurry of activity and the majority of the classic small molecule fluorophores, such as rhodamines (Figure...

...post we’ll provide an overview of some CRISPR mammalian expression systems, the typical applications for...what CRISPR components you will need to deliver. Minimally, one or more sgRNAs and Cas9 are required 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 ...

... them 17-26% smaller than other Cas13s and a whopping 33% smaller than Cas9! Their small size makes then...specific editor in mammalian cells Not all Cas13d proteins are functional in mammalian cells, but Konermann... easy to package in low-capacity vectors! These small, but mighty type VI-D enzymes are the latest tools... does not occur when Cas13d is expressed in a mammalian system. Since Cas13d is functionally similar to...compared CasRx to multiple RNA regulating methods: small hairpin RNA interference, dCas9-mediated transcriptional...

...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...genomic deletions We were inspired to produce intrachromosomal deletions based on the experiments of Kim ...References: 1. Lee HJ, Kim E, Kim JS. Targeted chromosomal deletions in human cells using zinc finger nucleases...palindromic repeats (CRISPR)/Cas9 nuclease system in mammalian cells. J.Biol.Chem. 2014 Aug 1;289(31):21312-21324... mammalian cell...

...to the XFP in the construct, dBrainbow includes small epitopes fused to each XFP. Secondary antibodies...signal. Imaging endogenous fluorescence in live animals only permits visualization of two colors. Similarly...protein microinjection or crossing of Zebrabow animals to an inducible Cre line. The Schier group developed...cell populations using the Confetti mouse For mammalian systems, the Clevers lab developed the Confetti...expression in the Ubow mouse Another murine reporter animal, the Ubow mouse (Ghigo et al., 2013), was developed...RFP are not present at the same frequencies. The small population size of CFP++ cells permitted researchers...

... to try to determine whether simpler and maybe smaller Cas systems could exist in nature. To do this,... Institute for uncharacterized genes that were proximal to a CRISPR array and the universal CRISPR integrase...cas4, and a new gene cas14. cas14 encodes for a small Cas protein (40-70 kDa), which is half the size ...

...virus- a non-integrating RNA virus Episomal vectors- extrachromosomally maintained DNA plasmids that express...Reprogramming efficiency of PBMCs with episomal vectors Episomal vectors are based on the Epstein-Barr ... pCXLE episomal vector system generated by the group is an optimized mixture of three episomal plasmids...expression from the other episomal plasmids. When used on PBMCs, the pCXLE episomal vector system increases...reprogramming purposes. In this post, I’ll describe an episomal vector system that increases reprogramming efficiency...thereby increasing the costs of the procedure. Episomal reprogramming is an ideal and cost-effective method...encoding reprogramming factors behave as separate extrachromosomal elements in the nucleus of the target cell...