...The importance of psychrophilic enzymes From an industrial perspective, the enzymes obtained from psychrophiles...diverse array of processes. The food processing industry, for instance, employs psychrophilic enzymes for...extremely useful benefits for any food processing industry as it can potentially help minimize the incidental...host The expression of recombinant proteins for industrial or research purposes requires stable host cells...either die out or grow so inefficiently that no industrial operation would ever consider them worth the ...continuously engaged with a variety of stakeholders from industrial and commercial operators, government agencies...iGEM teams to interact with personnel from the industry and brainstorm with each other. The event also...

...transfer buffer.  Adjusting the buffer For wet and semi-dry transfers, you can adjust the amounts of alcohol...you’re seeing odd background patterns, rather than just high background, your issue is likely technical....examples, like BioRad’s Western Blot Doctor page, or robust discussions, like ResearchGate’s western blot forum...calculations will not be correct, and you’ll need to adjust ‌your antibody dilution to accurately measure these...the detectable range of that antibody dilution. Adjust the antibody dilution until your protein dilutions...or low signal during the imaging process, try adjusting your exposure time first, since you can do that...protein amounts directly, stripping and reprobing. Just make sure to go from least to most abundant protein...

...for your protein of interest. CRISPR-Cas9 and a custom gRNA is all you need to generate a frameshift mutation...vector and Lenti expression vector. Turn it on for just a little while Yes, there’s a CRISPR for that, too... fused to a dead Cas9. The system is led, via a custom gRNA, to your gene of interest to temporarily increase...system requires tunable control - or maybe you’re just a control freak - this section is for you! Many ...lot of overlap with the systems discussed above, just with more elements of control. Tet promoters Tetracycline-dependent...Destabilizing domains (DD) are akin to degron tags, just with a flipped mechanism. When a protein is tagged... will bind and stabilize the DD, resulting in sustained protein levels. Looking for a DD? Check out our...

...available PAM must be selected. When it comes to CRISPR experiments, therefore, you often must make a choice...gene editing in research, the old PAM constraints just won’t cut it anymore. Say hello to PAM-free and ...top priority: location or efficiency. If you are just trying to shred a long non-coding RNA, cutting most...edit is reduced by over 50% when the cut site is just 10 bps away from the site in question (Kwart et ...necessarily more flexible than NGG; they’re often just different (e.g. NGAG, NGAN, etc.). Additionally,...solution to the issue. Nor are all Cas enzymes equal; just because an enzyme recognizes a PAM sequence in your...it. The SpRY Cas9 can also work with NYN PAMs; it just isn’t always ideal. But the increased flexibility...

...beyond the laboratory and seriously damage public trust in scientists, experiments involving potential gene...easy-to-implement confinement strategies that are robust to human error. Used in combination with standard..., and national authorities) was to detail these robust and easy-to-implement confinement strategies2. ...that seldom reproduce sexually because the drive must be much more efficient and minimally harmful in ...popular support, which in turn depends on widespread trust in scientists to develop new technologies with care... care and humility. Nothing would damage that trust more than an accidental gene drive release, which ...positive that at least some scientists could not be trusted to handle such a powerful technology responsibly...

...science policy, public outreach, and restorative justice.  Follow her on twitter @sung_vanessa. Albany, ...policies—many of which impact reproductive rights and justice. Whether it’s used to fight the myths that abortions...scientific community as we use for our science. "We must ask which perspectives and bodies are not present...celebration of science, at a time when scientists must rise up and become more politically engaged. I was...encounter hostile anti-science activists. However, I must say that I was supremely proud of Nashville that...promote harmful policies that benefit polluting industries.In March, three climate scientists and an economist...partisan: everyone (of every political persuasion) must be humble in the face of evidence. Stephanie Hays...

...employment within industry or academia; although the ACS career fair is more industry-focused. For another...what specifically should you do? What follows are just a few suggestions based on what I have found to ...ASCB) or the American Chemical Society (ACS) are just 2 examples of organizations that hold large, annual...another industry-geared option, try the Biospace career events page. Biospace holds career events all over...fairs is a great start, but you need to do more than just show up. Do your homework and know who is going ...contact an email with the relevant info and they just might personally hand your credentials to HR, greatly... for success in the career you hope to have, you must think outside of the box (most likely your lab, ...

...journals and several funding agencies (e.g. Wellcome Trust, NIH Data Sharing Policies, etc.) and is founded...founded on the principle that scientific publications must contain enough information (or links to information...anonymous request.  This means that supporting datasets must be logically annotated and arranged, as in an actual...services such as SlideShare, Figshare and Dataverse. Just as Addgene helps with DNA reagents, these resources...interpretation of data, publishing complete data, rather than just a single projection of it, could allow idea generation... hiding behind our jargon and esoterica. Public trust in and funding of science is not served when it ...attributes should be welcomed. However, expectations must also be reasonable, sensitive to the data type and...

...person, builds a strong sense of trust between coworkers. Since I trusted my coworkers so much, I knew that...the culture at Addgene cultivates trust between coworkers and that trust then translated into this incredible...bigger projects I was tasked with was creating and sustaining a biweekly report of our social media analytics... items, big takeaways, and timing. The sense of trust and comfort I had with my teammates acted as my ...enjoys reading, petting dogs, and winning the office Just Dance competition.    Special thanks to my manager...

...for differences in protein expression. (If you're just getting started, check out our Introduction to PCR...some general helpful principles (and I recommend Bustin, 2017 for a more in-depth review.)  Your amplicon... of primers should be between 100 and 400 nm and must be optimized experimentally.  Probe should have ...treatments may affect the assay. It’s unusual to need to adjust amounts of reagents in your chemistry, unless running...compared to your control. But as we said previously, just because it’s used often doesn’t mean it’s right ...difference (RQ>1), which may be easier to conceptualize. Just remember to switch them back before reporting the... flattery - imitation! Use the MIQE guidelines (Bustin, 2009) to ensure you’re recording and reporting...

...them. The adjustment is done within publication years. What do we find once we do this adjustment? That papers...Addgene? Probably not all of it. Our journal adjustment is just a first step. There are lots of other factors...papers with Addgene plasmids and those without, we adjust our estimates based on the journal that published..., methodologies, or qualities of research our adjustment controls for them.   So, for example, in 2010...Addgene plasmids were. Thus, to get our journal-adjusted citation measure we re-weight the 3 percent of...

...about [RNAi], ... it was just this complete black box. The RNAi process had just been discovered, the mechanism...case of CRISPR, the hype was completely justified… it has been just an incredibly useful technology.” He...several short talks by current faculty members and it just blew me away….  I have to come here, I need to spend...model at the Broad Institute because they are “not just developing technology for its own sake,” but work... do with CRISPR technology...” and “with CRISPR, just the results are so much cleaner. The time frame ...

...is imperative that you resist the temptation to “just do it myself” or micromanage –you don’t have time...required to develop the web page. The team manager must then facilitate appropriate communication between...Remember that there are always things that the manager must do. You can’t delegate the giving of feedback or...You can’t delegate tasks assigned to you and you must usually take on all tasks that involve confidential...regular check-in meeting on the calendar even if it is just 15 minutes a week. Clearly identify constraints...right away? Provide adequate support. The manager must be available to answer questions. If you are perceived...even be the best way. This facilitates success and trust. Micromanaging is bad for everyone. Document action...

...non-homologous end-joining (NHEJ) repair pathway is not very robust. In many cases, homology-directed repair does not... due to its condensed design. For both systems, custom spacers/gRNAs are easily inserted using BbsI and... from Streptococcus thermophilus, Cas9sth1 as a robust and highly active enzyme for transcriptional silencing...CRISPRi section, the Marraffini Lab published a robust tool for CRISPRi and CRISPRa using RNAP-Omega-dCas9...could be applied to diseases where the presence of just a single organism, such as Clostridium difficile... systems have been used successfully in mice for just such a purpose (Selle et al., 2020).  Targeting ...pair window of the target sequence which could be adjusted by changing the sgRNA length. The system could...

...also doesn’t require a ligation step. Based on the robust system of homologous recombination found in E. ...together through transformation, so the cloned vector must be ligated. But in LIC, longer single-stranded overhangs... by its sequence constraints. The long overhangs must not contain the dNTP present in the exonuclease ...recombination. SLIC inserts and linearized vectors must have homologous DNA sequences (20–60 bp) on either...matter if those overhangs perfectly match up. They just need enough complementary ssDNA to hold the plasmid.... The resulting recombination intermediate won’t just have nicks: it may have gaps or overlapping regions...dependence on single-stranded overhangs. These overhangs must be accessible to allow for complementary base pairing...

...way! Many thanks to guest blogger Kaustubh Kishor Jadhav! Kaustubh Kishor Jadhav, MSc is a Research Assistant...This post was contributed by guest blogger, Kaustubh Kishor Jadhav, a Research Assistant at MGMs Institute...lowered pH due to microbial contamination. Photo by Kaustubh Kishor Jadhav. Mycoplasmas can remain in the...the mycoplasma DNA appears in the form of small clusters, apart from the cellular DNA. Fluorescent DNA ...develop resistance to the antibiotics. Antibiotics must only be used to remove mycoplasma and not for preventing...CO2 incubators, used for maintaining cell lines, must not be overcrowded (filled no more than 60% of its..., reagent bottles and other sources of infection must be cleaned regularly and checked for contamination...

...the Advocacy Committee at the NPA for 5 years and just recently stepped down. I have gained invaluable ...deadlines and performing the work I agreed upon (just as I would in my laboratory work). Committee members...formats but I learnt to overcome the personal frustration and submit reports in the format requested which...alignment with the overall mission of the NPA. You must accommodate many different types of people in order...professional as I represented the organization, not just myself as a scientist. I learnt to wear different...funded by supporters such as the Burroughs Wellcome Trust. These “leadership institute’ weekends would entail...difficult and you can find opportunities around you. You just need to be prepared to take on the challenges and...

...information must be controlled independently; that is, the signal to record one bit of information must not act...vectors for customized lentivirus production Finding a lentiviral vector backbone with just the right combination... to create a tool that is quick, versatile, and robust. Dubbed the LACE system for light-activated CRISPR-Cas9...Toolkit for constructing fusion proteins with customizable N- and C-terminal modules   Thanks to the lab...Technologies. The MultiSite Gateway Kit is a customizable Gateway®-based toolkit that allows scientists...interest: Buj et al., BMC Molecular Biology. 2013 August 20th; 14(1). TRICK: A method for visualizing ...LacZ, puromycin or CreERT2 can be combined with custom-cloned vectors in a Multisite Gateway reaction ...

...designed with environmental sustainability in mind (Table 2). Table 2: Sustainability features highlighted in... large amounts of high quality DNA. Figure 1 illustrates the three grades of DNA you may need based on...their input (Table 1). When you feel like you need just a bit higher DNA yield, but not enough to go up ...getting sufficient plasmid DNA, you may need to adjust your starting culture conditions rather than your...impact, check out Addgene’s blog post on bringing sustainable practices for the lab. Plastic consumables (including...by more than half 70% recycled materials Sustainable forestry certification Starch-based compostable...Plasmids 101: Optimizing Plasmid Yields Bringing Sustainable Practices to the Lab: Easy Action Items Additional...

...folded ortholog that cleaved RNA robustly, unlike LwaCas13a, which must be stabilized by monomeric superfolded...quickly becoming major players in the CRISPR field. Just a year after Feng Zhang’s lab identified Cas13a ...an RNA nonfunctional. In their quest to build a robust RNA editor, the lab started with the Cas13 scaffold...ADAR constructs, like ADAR2DD(E488Q). They also adjusted the structure of the guide RNA by introducing ...that position. PspCas13b-ADAR2DD(E488Q) displayed robust editing with various spacer lengths from 30-84 ...editing from the Cas13b REPAIR system To test the robustness of REPAIRv1, the lab targeted 34 pathogenic G...