...to be low copy number. Conversely, plasmids that can initiate DNA replication independently are said to...AT-rich regions of DNA. In a test tube, the DNA helix can be separated by DnaA alone, as long as there are ...occupied by ATP-bound DnaA, the proteins cooperatively can separate the DNA strands. Created with BioRender.... available to bind DnaA. Fun fact! As far as we can tell, DnaA doesn’t need to hydrolyze ATP to do its...from the population, while replicating too quickly can slow cell growth or even kill the host cell. For ...replication of pSC101 depends on host-encoded proteins, it can be regulated indirectly via the same mechanisms that... to keep replication in check, high copy numbers can allow RepA to link iterons from two different plasmids...

...results in two DSBs that can be repaired by NHEJ restoring mCherry expression. Cells can then be sorted using...editing event. GFP positive cells can be sorted and have a significantly higher frequency of base edits ... created an array of base editing reporters that can help you do just that. Monitoring APOBEC and Cas9...due to a frameshift. Restoration of fluorescence can only occur through editing by a APOBEC-Cas9n-UGI ...Fluorescence-activated cell sorting (FACS) and the editing efficiency can be easily determined by comparing cells expressing...editing event would result in GFP fluorescence that can be visualized and sorted via flow cytometry. ...mutated start codon to initiate protein expression, it can be used for an array of detectable proteins including...

... functionalities, it can be difficult to figure out all the ways a given ELN can make your lab life easier... full activity on a project. Can you meet the ELN support team? We cannot stress enough how important ... It is important to know in which ways your data can be exported or printed from the ELN when needed. ... reporting on a project – printing your data out can be required every once in a while, so it is good ...your team. Do you need to make sure your research can be patented? I.e. is automated time stamping (detailed...the times and dates of your actions. These records can take the form of written comments, added files, notes...easier. If you can talk to an ELN support team directly, they should be able to walk you through your ...

... recombinant protein expression. Many challenges can arise when over-expressing a foreign protein in E...restriction/methylation system. This means the strain can neither restrict nor methylate DNA. dcm: Similarly...works well for most experiments, the lac promoter can “leak,” meaning that a low level of expression exists...mostly a problem for toxic protein products, which can prevent the culture from reaching the desired density...any) protein expression? Low expression outcomes can result from variety of sources, so fear not—there...not—there are a few simple troubleshooting measures that can help get you back on track: Compatibility: Double-check...Growth Media: Changing media is tricky, because there can be a trade-off between growth rate and protein quality...

...information. Our agility as a nonprofit resource can prove to be vital during this period of constrained...isn't lost. Universities and research institutions can turn to Addgene as a trusted partner for archiving... difficult to maintain or access. These services can be listed as direct costs from a CRO in grant applications...commitment to collaboration and resource sharing, we can navigate this difficult period together. The future...

...COVID-19 and coronavirus collection page, where you can find plasmid tools to accelerate your COVID-19 research.... Download the blue plasmid background.     You can find all of the plasmids that Addgene offers by visiting... visiting our collections page. Here, you can browse plasmids by field, organism, research technique, ...from Addgene are shipped as bacterial stabs? You can learn about what format your material will come as...

...this post, I’ll describe how rabies virus (RABV) can be used in the brain to visualize how neurons are... this, cell type specificity of the starter cell can be achieved by using EnvA-pseudotyped RABV, and only...in user-defined cells. For example, a researcher can deliver Cre-dependent TVA to Cre-expressing neurons...it is supplied in trans. Monosynaptic RABV spread can be achieved by modifying the wild-type RABV genome...Wickersham et al., 2007). Thus, RABV transsynaptic spread can be controlled by selectively supplying the G protein... Once in first order neurons, the G-deleted RABV can divide and replicate (since these processes are mediated...RABV monosynaptic tracing is a powerful tool that can be used to reveal brain circuitry, and continue to...

...mediate A•T-to-G•C base changes (Figure 1), but it can be challenging to make these base changes, especially... in action they bind DNA so that the TadA domain can edit the short single-stranded DNA stretch that is...catalytically impaired D10A nickase from ABE7.10 can be a problem. Therefore, the authors created ABE8...protospacer adjacent motif (PAM) made up of 5′-NGG-3′ (“N” can be any nucleotide base, followed by 2 guanines). ... in primary human cells  Cell therapy approaches can use base editors to correct mutations in stem cells...which cause sickle cell disease and 𝛽-thalassemia, can be mitigated with the expression of fetal hemoglobin...plays a role for off-target editing effects. ABEs can be introduced into cells as a plasmid or as mRNA ...

...provides the widest tropism or range of cells a virus can infect. Note: While both lentiviruses and γ-retroviruses...modify tropism. Briefly, different envelope plasmids can direct the production of virus with various tropisms...HIV for lentivirus). Additionally, γ-retroviruses can only transduce dividing cells because they are only... breakdown of the nuclear envelope. Lentiviruses can transduce both dividing and non-dividing cells because...plasmids or the packaging cell line. γ-Retroviruses can transduce target cells, but the transduced target...Are γ-retroviruses self-inactivating (SIN)? They can be. Plasmids that encode SIN γ-retrovirus lack most...widespread use of γ-retroviruses (gamma-retroviruses) in cancer and stem cell research has prompted the development...

...More importantly, you can use these four steps for any type of writing. You can use them to help you write...accessibility is removing barriers when we can and lowering them when we can’t. We know barriers exist in scientific...and after ultracentrifugation so that a scientist can extract virus from the correct layer.  Third, think...Where is the reader accessing the information? This can be physical or digital. For instance, you would present...results. And many journals accept alt text, so you can submit it with your manuscript and help make your...

...publication describing how short term storage on dry ice can shift sample pH, we wondered whether or not the dry...affected by dry ice, and specifically how dry ice can affect virus from Addgene. Bottom line: there’s ...temperatures (-80°C) and once in the tube, the CO2 can alter the pH of the liquid sample. The good news ... that this effect is reversible and the pH shift can be prevented. Keep this information in mind if you...C. For samples that are shipped on dry ice, this can affect the pH of the sample, which may have some ...Additional Resources on the Addgene Blog Learn How You Can Use the Viral Service Tips for 1st Time CRIPSR User...

...trade-offs. This balance can be extremely tricky to find: each of the factors can negatively impact both...or chemically reactive side chains, all of which can help determine structure and function relationships...relationship between structure and function…if you can get your protocol to work. Or, to quote Ryan Mehl...robust and universal techniques that researchers can easily implement in their own labs. They provide ...bulletin board on their website specifically so people can talk about GCE. In fact, their first protocol is ...proteins, you’re probably quite interested in the canonical 20 amino acids. But in your quest to learn more...code expansion (GCE): techniques in which non-canonical amino acids (ncAAs, also sometimes called unnatural...

... indicator. Archon1 can embed itself into the cell membrane where cell voltage can then be measured. Archon1...rapidly screen thousands of proteins for one that can report electrical activity via imaging. They took...nature, but developing them into optogenetic tools can be difficult because this requires detailed structural...interactions in cells because the protein binders can be customized for protein visualization, relocation...these photo-reversible protein-protein interactions can control subcellular localization processes in vivo...

...places they can’t reach on their own. The antibody’s heavy-chain constant domains, or Fc region, can be bound...isotypes. Binding to complement system proteins, which can initiate a cascade that helps attack extracellular...useful to know the isotype of an antibody so you can select an appropriate isotype control. An isotype...There are two IgA subclasses: IgA1 and IgA2.  You can learn more about isotype classes in our Antibodies... constant domain gene cluster. DNA recombination can delete intervening heavy-chain constant domain genes... antibodies Read about other techniques that you can use antibodies for in our Intro to Antibodies post...

...that lays out which experiments can be found in each folder. This can be a simple .txt document. Record...addition to output files, this is also where you can store any power point, excel, or other files relevant...the camelCaseFormat since spaces between each word can make it difficult to accurately call a file. Version... track of your exact changes. Alternatively, you can simply make a note in your methods section about ... Lab Notebook Organized Learn about Software You Can Use to Save Time and Track Your Experiments Try out...

...clone can then be monitored over time by sequencing the barcodes in the population (all barcodes can be ...of cells with specifc, easily read sequences that can be used to measure their clonal abundance before ...tumor xenograft models. She hopes that the library can be used to identify appropriate, non-overlapping ...development - such that initial resistant populations can be eradicated up-front quickly. This approach is ...25849130. 2. Hata, Aaron N., et al. "Tumor cells can follow distinct evolutionary paths to become resistant...resistant cancer cells exist prior to drug treatment. They infected a non-small cell lung cancer cell line...tool to look at a variety of questions surrounding cancer development, metastasis, and drug resistance, Bhang...

...smegmatis (Yan et al., 2020). NHEJ can also be engaged by variants of the canonical Cas9 approach. A pair of CRISPR...system for genome engineering is the fact that Cas9 can be easily programmed to make a DNA double strand ...frameshift mutation. Of some importance, the deletion can be less heterogeneous when constrained by sequence...loading of Ku and normal NHEJ activity. Other factors can also influence NHEJ activity. For example, repressing... flank regions of hundreds of base pairs or more can simultaneously introduce a pair of chromosome breaks...containing compatible overhangs is available. Cas9 can also be altered to generate a targeted single strand...and Biophysics and the Lineberger Comprehensive Cancer Center at UNC Chapel Hill.   References Bennardo...

... of the AAV we have released so far in 2019. You can also browse our entire AAV inventory! Our new AAVs...fluorescence of the GCaMP7 family of calcium sensors and can be used for imaging of small neuronal processes like...Since it’s illuminated by red/orange light, Archon1 can also be used in conjunction with optogenetic activators...AAV These fluorescent protein expression vectors can be used to visualize cells and to quantify targeting...expression to GABAergic interneurons. These vectors can also be used as the control vectors for experiments...

...Additionally, you can upload your own images to the Biorender workspace. Free Biorender accounts can store 5 workspaces...color and size of icons are adjustable. New icons can be requested, although there’s no guarantee Biorender...informal settings like lab meeting presentations. You can download larger images without the watermark, which...then clicking the “Usage rights” dropdown menu, you can search for images “Labeled for reuse with modification...Google Image Search results includes images that can’t be reused without attribution, despite filtering...

...victoria the functionalized nanobodies described here can be used to target any protein fused with GFP, YFP...vicinity with biotin. The biotin-labeled proteins can then be isolated and analysed. In this work, the ...School have developed a group of plasmids (here) that can be used to label all proteins within a radius of ...Tyrosine) with biotin. These biotin-labeled proteins can them be analyzed by mass spectrometry allowing a ...composition of proteins found at the telomere, and hope it can be used to expand our understanding of proteome localization...MarathonRT, isolated from Eubacterium rectale, significantly outperforms other RTs in this regard and is ...