...emotions, such as frustration or anger, because your discussion will get side-tracked, and you might create... your project and work hours. While specific strategies to deal with each of these personality types ...these principles it would be “assertiveness.” 7 strategies to become more assertive & resolve conflicts ...supervisors are difficult people. The following seven strategies will help you to become more assertive and resolve...between you and your supervisor, you need to demonstrate that they can count on you to follow through ...professional conflicts. At the workplace, these strategies will help you to communicate with confidence ...your supervisor is a difficult person, the above strategies will still be helpful but you will need to be...

...the E. coli B strain. Common E. coli strains used in the lab Most of the commercial strains you find today... the intestinal tract of animals. There are many different naturally occurring strains of E. coli, some...commercial lab strains of E. coli used today are descended from two individual isolates, the K-12 strain and the...the B strain. K-12 was isolated from a patient in 1920 and eventually led to the common lab strains MG1655...1918 but was first referred to as “B strain” in 1942. The BL21 strain and derivatives are the most common...changes found in E. coli strains. Table 1: Common gene mutations found in E.coli strains Gene(s) Description...of the popular strains, their genotypes, and their primary use in the lab. These strains are all based ...

...terminators and resistance cassettes for selection of transgenic plants. The complete GreenGate toolkit, now available...the time required to generate multi-construct transgenic plants. In the most practical terms, what GreenGate...comparison to more tedious and time-consuming traditional cloning methods, Lohmann said. The key component...like other endonucleases except that they cut at arbitrary sequences a defined distance away from the recognition...principle work in any plant species. Tricks of the trade Lohmann does have one special tip for anyone looking...efficiency, simply stopped working. They finally traced the problem back to the endonuclease. “It’s just...about it but be gentle. If it no longer works, just trash it.” Lohmann and his colleagues including Joachim...

...lab. Extracurricular activities don't have to be ‘extra’ Today’s graduate and postdoc training focuses...using multidisciplinary, collaborative, and translational approaches for scientific tool and technique...However, careers outside of the lab highlight “transferable” skills such as leadership, management, and ...in a myriad of professions is to engage in extracurricular activities (EA). EAs can: Introduce new perspectives...may expose which experiments are crucial in demonstrating the true innovation, impact and, in turn, the...opportunity to expand associations outside of the traditional academic setting and to shape and strengthen ...strengthen interests and skills. Although the word “extracurricular” may be taboo in the lab, multiple research...

...emission spectra for BV421 and BV711 on a 3 laser spectral flow cytometer   Using spectral flow When...autofluorescence extraction feature. Although autofluorescence extraction is an optional feature of spectral flow ...portion of that spectra to use for compensation.     Figure 1: The emission spectra and filters ...cytometer.   Spectral flow cytometry Full spectrum flow cytometry, or “spectral flow” cytometry, ... be distinguished using spectral flow since the machine uses the full spectral signature, not just a narrow...in the spectra can be used to distinguish these two fluorophores from one another during spectral unmixing...with spectral flow—you can run a three color panel or a 40 color panel! When preparing a spectral flow...

...Enhancer traps Like promoter traps, enhancer traps lack enhancer elements and require the trapping of endogenous...circumstances, trap methods may be the preferred screening option. Promoter traps Promoter trap systems lack... enhancer traps! However, don’t count the other traps out just yet. Gene and promoter traps unambiguously... up on their modern counterparts. What are traps? Traps – gene, enhancer, and promoter – all rely on...in fly, mouse, and yeast models. Why traps are useful Traditional genetic screens employ various on/off...expression of the gene they monitor. Gene traps Gene traps are promotor-less constructs that contain ...gene product resulting in a transcriptional fusion, much like promoter traps. While this system does not...

...an article describing a sucrose supplementation strategy to improve yield. In addition, as described in..., the team developed a viral genome sequencing strategy to confirm the identity and purity of all AAV ...! AAV purification by iodixanol gradient ultracentrifugation This video goes over AAV vector purification...protocol page for more detailed instructions. AAV titration by qPCR using SYBR green technology Here’s some...used to generate a standard curve, visit the AAV Titration by qPCR written protocol page. Getting started...

...competitive transplant assay in the mouse intestine, demonstrating its utility in long-term tracking in vivo...provides a combinatorial indexing strategy that simultaneously tracks clonal history and cell identity ...linking ancestral gene expression patterns with reprogramming outcomes (Biddy et al., 2018). Tracking lineage... tool that combines scRNA-seq and lineage tracking to track the dynamics of gene expression and clonal... of CellTag The Morris Lab developed CellTag to track the direct reprogramming of mouse embryonic fibroblasts...reprogrammed into iEPs by retroviral expression of transcription factors Foxa1 and Hnf4α, a process that like...reprogramming (CellTagMEF), at the end of the three-day transcription factor reprogramming treatment (CellTagD3), ...

...NEB’s new teaching space at Lab Central to learn all about it. LabCentral is a lab and office space shared...because both LabCentral and Addgene aim to make scientific advancement easier. NEBNext Ultra FS DNA Library... prior NEBNext Ultra II DNA Library Prep Kit for Illumina, one was only able to extract libraries from...New England Biolabs (NEB) hosted workshop at Lab Central and discuss why these types of workshops could ...in a lab.   Participating in a workshop at lab central We’re working on a project where we aim to do large...throughput DNA prepping by trying out NEB’s new NEBNext Ultra II FS DNA Library Prep Kit for Illumina sequencing...people to take a break and relax. Companies at LabCentral get access to a variety of experimental services...

...your AAV packaging transfection. In my lab we regularly use the ratios below: 1 Transfer Plasmid : 1 pHelper...prevent future headaches. To help you plan PEI transfections for AAV packaging, considering using this PEI...Calculator. AAV packaging typically requires transfecting three plasmids at specific molar ratios. To ...PEI calculator for AAV packaging Use the PEI transfection calculator for AAV packaging Select the plasmid...pHelper: 1 RepCap OR 3 Transfer Plasmid : 5 pHelper: 2 RepCap Select what type of tissue culture dish ...dish you will be performing the transfection in: 15 cm dishes OR CF5 stacks Select how many dishes or ...or stacks you plan to transfect: 1 to 10 Fill in experimental details on the spreadsheet: Name your...

... don’t wait. Start communicating now. These 9 strategies will get you started:  Subscribe to Career Blog...internships to cover their annual meetings. 8) Training – Get a formal education Some people get into ...journalism graduate schools where you may take a science track. If you don’t want to commit time and money to a...engagement for institutions and the Royal Society gives training in media skills, writing about your research and...learning courses at the National Council for the Training of Journalists (NCTJ) or the Alan Alda Center ...

...contain plasmids deposited at Addgene from the abstract page. Check out that beautiful Addgene “Find Plasmids... Plasmids” badge in the image below. If an abstract on PubMed has the “Find Plasmids” badge, you’ll know... your PubMed account and are browsing article abstracts you should see the “Find Plasmids” box on papers...

...CRISPR-associated Transposase) by the Zhang group. Mechanism of CRISPR-transposons One of the CRISPR-transposon systems...DNA transposons have been hindered by their limited programmability: depending on the transposases involved...-guided transposition. By natively combining the DNA integration capabilities of transposases and the ...crRNA.  Three transposase proteins (TnsA, TnsB, TnsC). The donor DNA (aka mini-transposon), which contains... each type of CRISPR-Transposon system. (B) Mechanism of RNA-guided transposition in type I-F INTEGRATE.... Separately, TnsABC forms a transposase complex with the mini-transposon, excises it from the donor molecule...characteristic of the Tn7-family of transposons. Insertion of the mini-transposon can occur in two different orientations...

...way to begin the search is by using selection strategies, where only cells that have gained or lost a ...plasmid cloning.  But oftentimes, a selection strategy is not enough on its own to help you find your...plasmid. In many cases, you will need a screening strategy as well. In a screen, you aren’t killing off a..., then screen Why add a screen to your cloning strategy? A screen will help you more easily identify successful...information (more on this below). Plating the transformation on ampicillin plates containing X-gal allows...Fortunately, for the lab researcher, when the substrate X-gal is broken down by 𝛽-galactosidase, it turns...blue-white screening at Addgene! Some bacterial strains contain lacZ in their genome, and scientists have...

...repeats (ITRs) are what makes an AAV transfer plasmid an AAV transfer plasmid. ITR sequences are 145 bases...read through this challenging sequence. Transgene The transgene is the gene that will be delivered by the...Most AAV transfer plasmids have a promoter, but this element isn’t required when the transfer plasmid ...control transgene expression. Lox sites are directional 34 bp sequences that can flank the transgene and ...(WHP) Posttranscriptional Regulatory Element (WPRE) forms a tertiary structure when transcribed which ... nuclear export of RNA and RNA translation, and promotes RNA transcript longevity. The polyA is located...proteins needed for the virus to replicate,  and the transfer plasmid which contains the viral genome. In today...

...imaginations to use in this world where we are often trained to follow protocols, guidelines, and standards.... an odd or quirky lab story you’d like to see portrayed in LEGO form, feel free to share with us at BrickingScience...

...and Harvard Medical School. Scientists making transgenic organisms with Cas9 should be aware of the potential...conducted with extreme caution. Why does making transgenic organisms with Cas9 risk creating a gene drive... there are many easy-to-implement confinement strategies that are robust to human error. Used in combination...these robust and easy-to-implement confinement strategies2. Our hope was that anyone else who came up with...improve safety. What safeguards and confinement strategies are available? Molecular confinement involves...drives that can spread through populations of transgenic laboratory organisms but not wild organisms. ...targeting a synthetic sequence will only spread in transgenic laboratory populations with that sequence2. Both...

...serotype. Additionally, we check viral transgene expression in transduced cells whenever possible. This post... for transgene expression. Once packaged in AAV, this pre-recombined DNA could lead to transgene expression... users titrate their AAV vectors to find the optimal dose that allows for sufficient transgene expression...to make sure they are present at a very low concentration. So how do we make sure to catch all of these...particles (Figure 1). Briefly, packaged DNA is extracted from purified, DNase-treated AAV and submitted...with AAV-68544 to validate our method. We then extracted the DNA and processed the mixed sample in our ... research-grade materials. Addgene is going the extra mile and pioneering the systematic use of this new...

... Michael Davidson and Roger Tsien Commemorative Travel Awards. These awards are intended to help scientists...research with the academic community by funding their travel to a conference of interest. In this post, we’re... Michael Davidson and Roger Tsien Commemorative Travel Awardees: Xin Zhou and Fatima Enam. There were ...protein known to oligomerize when exposed to ultraviolet light. To do so, she generated fusion proteins...of an enzyme’s active site. In these fusions, ultraviolet light exposure caused oligomerization of the ...still block enzyme function upon exposure to ultraviolet light. Previous iterations of FLiPs only used...Cdc42, Raf1, Mek1, Mek2, Cdk5, and Cas9. Xin has demonstrated the utility of these FLiPs in a variety of research...

...vivo.  But what ultrasound lacked was reporter genes that could produce ultrasound contrast. That is, until...before imaging with ultrasound to observe gas vesicle-specific ultrasound contrast where the bacteria ...via regular intraperitoneal doxycycline injections to enable their tracking with ultrasound imaging (Fig...the transient cotransfection of the mARG plamsids into HEK293T cells using a chemical transfection reagent... scatter sound waves and thereby produce ultrasound contrast. We (the authors) are currently working on...also straightforward: after the E. coli have colonized tumors, inject L-arabinose intraperitoneally and...phase contrast microscopy, GVs visible in cells via TEM, and purified GVs under TEM. (c) Ultrasound imaging...