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... are equal. Created with BioRender.com. This kind of normalization is a nearly ubiquitous tool, but... assumptions that their loading controls are independent of these other sources of error. It’s important...housekeeping protein in your context. If you don’t find anything, you’ll need to conduct a control experiment...strongly influenced by unexpected variations in any individual protein. This ensures you are truly comparing...with β-actin (1:2000). Representative of three independent blots. B–C) Graphs showing the relative intensity...increasing concentration of β-actin in panel A. Indeed, the quantification in panel B is not especially...total protein are compared. While a higher slope indicates higher sensitivity, linearity is the most important...

...-4 colonies to ensure that you find the correct product. Once you find a colony with the appropriately...Posts Resources on Addgene.org Find Empty Vectors for Your Research Find Additional Protocols Browse Our... Some of these include, TA cloning, ligation independent cloning, TOPO cloning, one step cloning, and ...Li, et al., in 2011. This method is ligation independent, it does not require purification of insert or...simplified, purification-free, sequence-and ligation-independent PCR cloning method." BMC biotechnology 11.1 (...

...to be binding at particular genomic sites when in fact they are just non-specific or indirectly interacting...is the go-to method for mapping where a protein binds genome-wide, and has been widely applied in many..., an enzyme-based protocol for identifying DNA binding patterns that overcomes some of the limitations...Protein A fused MNase is then added and Protein A binds Immunoglobulin G (IgG) thus targeting MNase to antibody...provides an excellent additional measure of DNA binding sites in combination with existing ChIP-seq data...with CAPTURE Additional Resources on Addgene.org Find simple lab protocols Browse CRISPR plasmids for ...

...s-level job postings on a job search website like Indeed. Do the job descriptions sound interesting? Is ...work something you can see yourself enjoying?  To find master’s-level postings, try adding “master’s” to...filters to limit results to people that work in industries you’re interested in (i.e. biotechnology, pharmaceuticals...switch your field of work, both in academia and in industry. Advancing along a career path in these two fields...fields looks different however. In industry the career ladder is larger and there are more opportunities...Career advancement for PhDs, both in academia and industry, often means moving from the lab to a desk, although...on Addgene.org Check out Addgene's job openings Find eBooks from Addgene, including the Science Career...

... PhDs Making the Leap to Industry." He successfully transitioned to industry after his PhD and a couple...mental freedom. Practical approaches Engaging in mindful meditation, taking field trips, and spending time...gain fresh perspectives. Such activities foster a mindset that embraces the unexpected and encourages exploration...Serendipity is not just a stroke of luck; it's a mindset that can lead to unexpected discoveries and innovative...successful research career, whether within academia or industry. Albert Einstein once said: “There are only two...serendipity into your research but also cultivate a mindset that is adaptable, creative, and poised for groundbreaking...

...feedback when it is merited. Surprisingly, some people find this hard so get used to it by making it a concrete...However, I have also found that scientists seem to find it easier to give feedback in formal sessions using... Here are the 3 things that worked least well. Find out the Effect of the Feedback: How does this feedback...over, what would you do differently? Beware of indirect language You must be honest about performance ... mistakes I see is managers who hint at or use indirect language to talk about real performance issues...issues. Watch this little video to learn more about indirect language from Steven Pinker. As a manager you ...and clear. There is place in human discourse for indirect language, but it is not the way to go when giving...

...single peak, indicating one amplicon and no primer dimers. (b) A shoulder peak indicating primer dimers...the right conditions and parameters, some people find it can detect even smaller differences, and it can...dimers. (c) A double peak indicating 2 amplicons. Image credits (respectively): Zuzanna K. Filutowska; Selma...different types of pipettes and pipetting methods to find what’s most consistent in your hands.  When using...target in the reaction should be compared to an individually run sample when developing the protocol and ...and optimized until the multiplexed and individual values match across the board. If the samples or sample... e-book on applying MIQE guidelines! With these kinds of resources, you will soon become a qPCR master...

...short-hand for Clustered Regularly Interspaced Short Palindromic Repeats, in Pubmed returned 728 articles (3/12...because the Cas9 nickase doesn't have the ability to induce double-stranded breaks like the wildtype Cas9 does...'G'. Q4: Where are double strand breaks (DSBs) induced, compared to where the target site sequence (protospacer... colonies, then genotype them, it is possible to find both single-allelic and bi-allelic cells. Single-allelic...double nickase system, with PX335 vector, you need to find two guides for each cleavage site, but this system... we usually pick 3–6 guides around the region to find the most efficient guide (most guides would work...single-cell colonies, grow them up, then perform individual genotyping (using Sanger sequencing, for example...

... autophagic activity, and are a more reliable indicator of autophagic activity. The aim of this post is...emits red and green fluorescence. Following the induction of bulk autophagy, Rosella is targeted to the ...interest to track the autophagy of that protein. Find the cytosol-targeted and mitochondria-targeted Rosella...nm to 440 nm serves as a measure of autophagy induction: when Keima is in the cytosol, this ratio is low... this biosensor can’t be used with fixed cells. Find the mitochondria-targeted Keima variant at Addgene... in yellow puncta but a decrease in red puncta. Find the mRFP-GFP-LC3 plasmid at Addgene! GFP-LC3-RFP-LC3...fluorescence suggests a decrease in autophagic flux.   Find the GFP-LC3-RFP-LC3∆G plasmid at Addgene! References...

...for scientists working at home, and you will still find this useful. Now that we’ve been doing this for ...safe space. No matter why, now is a great time to find a few colleagues to work together on career and ... “the ability to assess and initiate things independently” and “the power or opportunity to act or take...format is good for everyone including those that find events stressful. Video chatting makes it pretty...effectively is a valuable skill. There are many different kinds of grants that you can apply for. If you are in ..., your PI is likely working on a number of them. Find one to apply to or offer to help with lab grant ...Opportunities Open Education Database: 100 Places to Find Funding For Your Research University of Florida:...

...still invested in seeing how they do it.    Finding Conflict in Science  In all science communication...conflict into your work, but one of my favorite is to find it within the science itself - within that process...trying to understand your completely unexpected findings, or understanding why your experiment has not ...not worked…again…for the seventh time….   I find that once I sit down and talk to a scientist about their...paper was actually from a completely unexpected finding in a different experiment, this tool was developed...fodder for science communication. The trick is to find one or two roadblocks that sparked creativity, changed... a story around them. A good rule of thumb is to find the pieces that make you think, “Oh, that’s interesting...

...student hasn’t been asked the question: “Academia or industry?” Once academia was the clear answer with a well-defined...web-based, career-planning platform called the Individual Development Plan (IDP) and uses it to match qualified... qualified scientists to jobs in industry, academia, and government. This great resource allows the undecided...and other professions increases your chances of finding a well-suited job. EAs shouldn't be viewed as an...interests: Use career planning tools such as the Individual Development Plan to generate a list of potential... more information Follow people in academia, industry, and government on social media (Twitter, Facebook...potential job Join LinkedIn and other job sites to find open positions and view their required qualifications...

...seamless manner, leaving no sequences or mutations behind. Furthermore, piggyBac™ offers a large cargo-carrying...cells lines, including stem cells, without leaving behind any accessory DNA sequences [2], [3]. Simply put...integration [4], the donor cassette is removed leaving behind no accessory transposon sequences.  The use of ... plasmid after excision by PBx, thereby leaving behind only those cells with a Footprint-Free™ genome ...this method can be effective, a loxP site is left behind in the genome following donor removal, which alters...somewhere between one hundred and one thousand clones to find a cell line with the edit of interest. With the ...Cathomen, and K. Schwarz, “Zinc-finger nuclease-induced gene repair with oligodeoxynucleotides: wanted ...

...targeting and sequencing millions of cells. For every individual sgRNA, you recover a minimum of 1,000 targeted... 5% off-target frequency is something to keep in mind, but likely will not have a significant impact on... many predictive online tools which can not only find gRNAs for you, but also score their probability ...probability of off-target events: CRISPOR, Cas-OFFinder, CCTop, and many more. While such software typically only...at mismatched guide sites, but not necessarily binding to these sites. Thus, these enzyme swaps will only...need to do your due diligence to ensure they are indeed a non-issue. Above we mentioned how to predict ...tested with some success. The major con is that binding by Cas does not always correlate with catalytic...

...expected fragments for that plasmid. This phenomenon indicates the presence of a multimer.   Fig. 1:...multimerization has been observed to occur in many different kinds of plasmids, taking many forms (Levene, 2009; Higgins... al., 2010; Cook et al., 2020). CGE uses laser-induced fluorescence (CGE-LIF) for detection and can provide...multimerization. One study by Bacolla et al. (2011) indicated that strain HB101 produced the highest yield of... at reducing multimerization and define recA-independent mechanisms. Avoid overgrowing the bacterial culture...-B DNA-forming sequences and WRN deficiency independently increase the frequency of base substitution ...26104344. Fishel RA, James AA, Kolodner R. recA-independent general genetic recombination of plasmids. Nature...

... are interested in understanding the mechanisms behind Mycobacterium tuberculosis resistance to pyrazinamide... and recombination-mediated genetic engineering (find more information on bacterial engineering using ...Devarshi Gajjar, The M. S. University of Baroda, India  Devarshi Gajjar has 10 years experience in the ...expressing fluorescent proteins will be used to tag individual strains of bacteria. Their interactions and growth...University of Ibadan, Nigeriaunder supervision of Dr. Akindele Oluwatosin Adeyi, University of Ibadan, Nigeria...PhD Candidate working under the supervision of Akindele Oluwatosin Adeyi at the University of Ibadan. ...regulation and diabetes will be expressed from plasmids (find more information on bacterial expression systems...

...found yourself frustrated reading through a paper to find that there is insufficient information about which...also designed to be persistent and exist online indefinitely. PIDs historically referred to objects that ...identify a plasmid available through Addgene and find verified information about each plasmid item. Given...associated plasmid page on the Addgene website and find specific information about the plasmid: name, description...and more. The plasmid page also helps researchers find other publications that have used the same plasmid... Addgene ID # and the information necessary to indicate that this ID is managed by Addgene. How to cite...down to the bottom of this plasmid’s page you will find that the “How to cite this plasmid” section now ...

...represent the fraction of that total with the indicated function. One huge reason CRISPR has become such... left the arduous task of virus production to individual labs, but now we’re very excited to provide ready-to-use...an entire lentiviral production protocol only to find that the low titer of her prep makes it unuseable...target these widely studied signalling pathways. Find Validated gRNAs to use with your Cas9 Virus Pooled...uses nearly 80,000 gRNAs to target around 20,000 individual genes. We provide enough lentivirus for you to... lentivirus production protocol here and you can find more tips and protocols for working with lentivirus...Viral Service Page Check Out Our CRISPR Guide Pages Find Viral Vectors ...

...size of the insert. You can search NCBI for YGOI to find the particular reference sequence if necessary. ...The plasmid was digested with 2 unique enzymes (HindIII and BamHI) and run on an agarose gel. The resulting... (*). When the plasmid is digested with either HindIII and BamHI alone (lanes 4-5), there is a single ...size of the plasmid. The double digest with both HindIII and BamHI (lane 3) produces bands at 6kb and 1.2kb...bromide (EtBr) to your gel before pouring it. EtBr binds to the DNA and allows you to visualize the DNA under...vist Addgene's resource for plasmid verification to find additional tips and detailed protocols on topics...Resources: Addgene's Molecular Biology Reference Find More Protocols and Videos      ...

...won't cut it for bioinformatics. All kinds of biologists are finding themselves using computational approaches...Additional Resources on Addgene.org Find Protocols for Your Next Experiment Find Our List of CRISPR gRNA Design...data sets (myself included) and we are faced with finding the best system to document these types of analyses...bioinformatics. In fact, there are endless ways and everyone finds their own. Here, I simply outline what works for...not apply or may not work for you. The best way to find a system that does is through trial and error, and...