...in the fields of gene and cellular therapies.     Additional Resources on the Addgene Blog Read What ...Technical Support Calls Resources at Addgene Visit the Addgene Careers Page Check Out Our Career Videos...You will also help troubleshoot products, give general experimental guidance, and resolve complaints about...willingness to help others with their problems, a general positive attitude, and an open-minded personality...will do well in a technical support role. What general characteristics do jobs in technical support have...covered by the lab based team as part of their general job duties. Bigger companies have dedicated technical...post. Klaus Wanisch is an Outreach Scientist at Addgene. These days he's doing a lot of science communication...

... terms of the vocabulary--DNA, protein, plasmid, gene, etc. Though I talked about CRISPR’s molecular mechanism...quick tips to get things going: If your PTA has a general meeting and a means to request funding for events...7th grade biology teacher. I led up to my job at Addgene, which is likely a career path none of them had...offered engaging examples of how understanding genetics is so timely.” Genome engineering, and many other...guidance and feedback. Additional resources on the Addgene blog Browse our CRISPR blog posts Download our...science communication blog posts Resources on Addgene.org Check out our comprehensive CRISPR guide Find...

...the expected complete sequence for the lacI/lacZ genes (1), while sequencing of white colonies showed a...PMCID: PMC6061786. Additional resources on the Addgene blog Learn the basics of Golden Gate assembly ...other plasmid cloning techniques Resources on Addgene.org Find basic molecular biology protocols Find ...

...T-cell development, physiology and malignancy." Genes & cancer 6.5-6 (2015): 184. PubMed PMID: 26124919...PMCID: PMC4482241. Additional Resources on the Addgene Blog Writing Scientific Manuscripts: Literature...Manuscript 6 Tips for Grant Writing Resources on Addgene.org Check out Our Hot Plasmids Articles to See Compiled...

...studies show that certain mutations of the BAP1 gene play a role in susceptibility to developing mesothelioma...Additional Resources on the Addgene Blog Find out More About Cancer and Addgene's Cancer Collection Pages...others never do, but we do know that there are some genetic factors involved. For example, multiple studies...research is being done to see what, if any, other genetic factors may be in play. However, there is no question...options: Early detection, immunotherapy, and angiogenesis inhibitors New research is also going into two...bevacizumab (Avastin). This drug (known as an angiogenesis inhibitor) prevents the growth of new blood ...the use of asbestos, thus ensuring that future generations will be protected from this dangerous material...

...delivery of genetic cargo  By Alyssa NeuhausTo improve delivery and reduce off-target effects of gene therapy...system to improve on-target delivery of genetic cargo Optogenetic proximity labeling and TransitID Tracking...continuous evolution and protein engineering to generate seven new prime editors, PE6a–g, that overcome...10.1038/s41467-023-40788-8. PMID 37612276.   Optogenetic proximity labeling and TransitID By Mike Lacy...transcriptional history of a cell. Lin et al. generated a fusion protein called iPAK4 that assembles into...43R] has been one of the hottest antibodies in Addgene’s collection since we began distribution and has...[K28/43R] under the Application section of the Addgene antibody page, and learn more about the whole-brain...

...my students published their work identifying new genes required for stem and progenitor cell maintenance...incorporated a discovery-based lab on planarian regeneration into one of my own courses, I was excited to...be authentic – students should be attempting to generate new knowledge that will be of genuine interest...Provost’s course recently used a combination of phylogenetic analysis, structural prediction, and bench work...recombinant enzymes in bacteria, purified them to near homogeneity using a 6XHis tag, and compared their specific...

...to make sense of biology, with a special focus on gene editing techniques. References and resources References... versatile genome manipulation. Nature Reviews Genetics, 24(3), Article 3. https://doi.org/10.1038/s41576...DeepSpCas9, a deep learning–based model with high generalization performance. Science Advances, 5(11), eaax9249...). Chromatin context-dependent regulation and epigenetic manipulation of prime editing. Cell. https://...j.cell.2023.03.034 Resources More resources on the Addgene blog How to Design Your gRNA for CRISPR Genome ...and Flexibility to CRISPR Editing Resources on Addgene.org CRISPR: PRIME Edit Collection The CRISPR Guide...

...from transient overexpression of protein fusions to gene-edited cells can decrease the number of fluorophores...Fluorescent Protein Should I Use? Resources on Addgene.org Generate Fluorescent Protein Fusions Request Plasmids...in-cell labeling strategies have two parts: (1) a genetically encoded “tag” expressed as a fusion with your... dye ligand (e.g., FlAsH, ReAsH) and a short genetically encoded tetracysteine (Cys4) peptide tag could...TMP-tag) – These widely used systems consist of a genetically encoded enzyme variant tag that reacts specifically...such as paclitaxel. Unlike other systems, no genetically encoded tag is required as the small-molecule...strategies have trade-offs between the size of the genetically encoded tag, the speed and selectivity of the...

...contain either an ampicillin or kanamycin resistance gene. Every step along the way involves good aseptic ...of tube for each plasmid we have at Addgene. Each sample at Addgene has four layers of location which can...pages! Resources Avaliable on the Addgene Blog Read more Inside Addgene Posts! Learn all about Plasmids ...Every plasmid sample enters Addgene the same way. A package is delivered by a mail courier and then the... a microcentrifuge tube. Once a sample enters Addgene’s lab, a series of events is triggered. Each step...fluctuates throughout the year (see graph below). Addgene scientists inspect each sample and make sure they...takes over and prepares the sample for entry into Addgene’s repository. For long-term storage in our facilities...

... Ti plasmids in Agrobacterium tumefaciens. Genes & Genetic Systems 77:1–9 . https://doi.org/10.1266/ggs... Resources on Addgene.org Download the Plasmids 101 eBook  Browse plasmids at Addgene Read our Molecular...plasmid partitioning system in the bacteria, but generally it consists of a centromere-like region on the...Kamruzzaman et al., 2017). Additional resources on the Addgene blog Read all of our Plasmids 101 blog posts Learn...

...likely -/-, 50% are -/+, and 25% are +/+ for your gene of interest. Thus, to find a null clone, you should...edited. For many experiments, the end goal is to generate a clonal cell line, so it’s important to verify... for a needle in a haystack before going on to generate and screen individual clones. If a clonal line...al., NAR). This technique uses the trace file generated in Sanger sequencing and a simple algorithm to...with your sgRNA sequence. The TIDE software will generate a graph representing all insertions and deletions... 4 clones. Did you use a dual guide system to generate your knockout? Then you’re in luck! You don’t ... to visualize an edit. Most dual guide systems generate a deletion large enough that it can be visualized...

...IPTG. T7 RNAP is then available to transcribe the gene of interest from a T7 promoter on the plasmid. Many...::T7 gene1 [lon] ompT gal sulA11 R(mcr-73::miniTn10--TetS)2 [dcm] R(zgb-210::Tn10--TetS) General protein...cytosine within a particular sequence. Browse Addgene's curated list of Bacterial Expression Systems Table...DE3) F- ompT lon hsdSB (rB- mB-) gal dcm (DE3) General protein expression BL21 (DE3) pLysS* Chloramphenicol...ompT lon hsdSB (rB- mB-) gal dcm rne131 (DE3) General expression; not recommended for toxic proteins ...lon hsdSB (rB- mB-) gal dcm araB::T7RNAP-tetA General protein expression  BLR (DE3) Tetracycline RecA-deficient...overexpression? The strains described above should generate sufficient expression levels for most purposes...

...mediated HDR) enables the tagging of endogenous genes close to native expression levels. However, it is...chimera are encoded by DNA. Since researchers can generate almost any DNA sequence in the way that they like...proteins is relatively straightforward. However, generating a fusion while keeping all of the native properties...challenging. In this blog I discuss strategies to generate fusion proteins and highlight some aspects of ... from Aequorea victoria and its variants are genetically encoded fluorescent probes. One of the limitations...restriction enzymes in the multiple cloning site for generating fusions. For the POI-fluorescent protein fusions...sufficient. For the fluorescent protein-POI fusions, we generate straight fusions, without any linker. When both...

...achieved through, for example, a previously induced gene knockout. Besides amine- and DNA-binding dyes, nucleoside... on Addgene.org Ready-to-use recombinant antibodies at Addgene Addgene's Antibody Guide Addgene's Antibody... combines a marker with a signal. For clarity, Addgene uses 'labeling' to refer to antibodies and 'staining...Find streptavidin plasmids and biotin plasmids at Addgene!     Figure 1: Indirect detection of target...signal. Dump gating When you are working with heterogeneous cell mixes, like bulk lympho-/splenocytes, you...Antibody Plasmid Collection Resources on the Addgene blog Antibodies 101: Reading a Flow Plot Antibodies...

...do not naturally express the protein or have had gene expression knocked down or knocked out. If you are...antibodies Resources on Addgene.org Find written and video protocols Visit Addgene's antibodies collection...plasmid encoding your protein of interest from Addgene, transfected it into your target cells, now what...wet, semi-dry, and dry but all follow the same general principle where the gel and membrane are “sandwiched...may even find plasmids encoding antibodies from Addgene that you can make on your own.  To narrow your ... found this blog helpful be sure to check out Addgene’s blog for overviews of other antibody topics. ...10.4103/1947-2714.100998 Additional resources on the Addgene blog Find other tips and tricks for molecular ...

...which come from naturally-occurring immunoglobulin genes, recombinant antibodies are plasmid-based. Their... What is an Antibody? Resources on Addgene.org: Browse Addgene’s Antibody Plasmid Collection Find antibodies...cascades that clear these infections. Anything that generates an antibody response in the immune system is referred...in the DNA of the hybridoma can occur, known as genetic drift, meaning that over time, the antibody you...monoclonal antibodies, recombinant antibodies are a homogeneous population. However, unlike monoclonal antibodies...antigen binding, the variable regions alone can be generated as a fusion protein. An ScFv is made up of the...detect the presence of a single protein from a heterogeneous mixture - for example, cell lysate or media ...

... 18 lung cancer patients. This study used CRISPR gene editing to knockout PD-1 on immune cells harvested... Cell heterogeneity Tumors are composed of a wide variety of cell types. This cell heterogeneity is an...cells that escape the elimination stage acquire genetic alterations and alter cell-surface antigen production... promotes the production of VEGF to increase angiogenesis around the tumor. This in turn regulates the... immunotherapy. Using activated DCs ex vivo to generate cytotoxic effector T cells, and targeting specific...played a huge role. For instance, researchers are genetically altering tumor cells and immune cells to knockout...element of successful tumor progression. A tumor’s genetic composition is constantly in flux and is dependent...

...procedure and application to the major ALS/FTD disease gene C9ORF72. eLife, 8, e48363. https://doi.org/10.7554...101: Validation Resources on addgene.org Addgene's Antibody Guide Addgene's Antibody Protocols... outlined below. Knockout cell line approach Genetic knockout-based validation can conclusively prove...you are working with cells that are amenable to genetic knockout, this represents a very robust approach...to start with a screening procedure in easily genetically modified cell types (top). The specific approach...10.1038/s41598-019-55133-7 More resources from the Addgene blog Introduction to Gating in Flow Cytometry Antibodies...

...transition from weak, random binding to precision gene editing requires three major steps: RNA loading,...resources on addgene.org  Addgene's CRISPR Guide Addgene's CRISPR 101 eBook  Resources on the Addgene blog Components...from Streptococcus pyogenes.   Figure 2:  Crystal structure of S. pyogenes Cas9 in the apo state...    Figure 3: Crystal structure of S. pyogenes Cas9 bound to guide RNA (PDB ID 4ZT0), shown opaque...sequence isn’t in place yet.     Figure 4: S. pyogenes Cas9 in the apo state (PDB ID 4CMP, top) compared... residues in the CTD can get cozy with DNA. S. pyogenes Cas9 recognizes the PAM sequence 5’-NGG-3’, where...complementary strand is mismatched.   Figure 5: S. pyogenes Cas9 bound to guide RNA and target DNA (PDB ID...