...several years ago we partnered with Seqwell to establish a next-generation sequencing (NGS) QC platform...including only titration of AAV particles and a protein stain to assess purity. For labs that produce a small ...samples, serotypes, or both. Unfortunately, the standard AAV QC regimen does not address this.  Previous...analysis approach can be easily incorporated into standard QC regimens and will provide scientists with the...

...we are cloning an ORF, we want to clone from the start codon (ATG) to the stop codon (TGA, in this example...give specificity and to also be compatible with a standard PCR reaction (see PCR Video). Therefore, our Forward...reverse complement to get PCR amplification. We can start similarly, taking the final 18 bases of the ORF,...restriction cloning post. However, even more than with standard restriction cloning, it is essential that you ...

...usage has been limited by technical problems. For instance, it’s much more difficult to introduce a gene ...primary (hESCs) and cultured cells showed up to 70% resistance to diphtheria toxin, indicating a 70% frequency... adapted from D’Astolfo et al. In certain instances, the potential benefits of nonviral transduction...protein purification (once a protocol has been established) may be less laborious than viral purification...

...a great start to a mentoring relationship, but if you don’t take this past the chatting stage you will...list available from this post. Use an activity to start a discussion. The Forced Choice Analysis is a fun...clarify and discuss values. The activity is just the start – it is the discussion after that will lead to learning...

...Standard AAV vectors are generated with a producer cell line like the widely used 293T. During vector...glioblastoma). They also found that exo-AAVs were more resistant to neutralizing anti-AAV antibodies. This is particularly...thought to enhance the production of exosomes or stabilize their structure or their interaction with AAVs...areas that are difficult to reach, and are more resistant to inactivation through anti-AAV antibodies. ...

...reagent is crucial for reproducibility because mistakes in this can lead to wasted time or retractions... the Addgene catalog. These IDs allow you to unmistakably identify a plasmid available through Addgene...all Addgene plasmids! In an effort to support standardized persistent identification of research reagents...Resource Identifiers (CURIEs) combined with well-established prefix resolution services such as Identifiers.org...

...is fine for its goal of creating a choice of standardized constructs and tools for the rational and simple...Golden Gate Assembly also allows the creation of standardized DNA constructs, but its use of Type IIS restriction...polymerase used in the amplification. This tool allows instant updates as fragments are added, deleted, or reordered... in biofuel generation, green chemistry and understanding the minimal genome. Many of these techniques...

... (Fig. 1). However, once thawed, there was a substantial reduction (from 7.4 to 5.0) in the pH of samples...antibodies, because of effects of pH on protein stability (Murphy et al., 2013). However, the pH changes...infectivity of the virus. However, there was a substantial reduction in infectivity after 3 weeks of exposure...der Geer P, Manning MC, Fitchmun MI. Protein instability following transport or storage on dry ice. Nat...

...range of characteristics (color, brightness, photostability, maturation, oligomerization), many of which...choosing a protein, we often direct them there as a starting point.  However, there is a great deal more information...database relating to that specific protein (for instance: EGFP).  Users can “favorite” certain proteins...of FPbase FPbase is still very much in the early stages of development, and there is plenty of room for...

...abundance, the eGFP will be stable and fluoresce. But if the variant is unstable, it will be degraded, leading... of fluorescent reporters. Bacteria containing unstable fluorescent proteins under the control of promoters... cell cycle and growth rate using TIMER and an unstable fluorescent protein reporter under the control...

...at odd times, new strategies can make socially distanced work more creative. An example could be making...signal the approach of a brain break, where everyone stands up, disappears from the video, and prepares a coffee...brainstorm on data in smaller groups, where for instance the presence of the PI in the main room inhibits...these weird times of shift working and social distancing. The take home message is the approaches/tools...

...give rise to a starfish shape. You can ask me about the different arms of this starfish sometime, but... plug-in called Maps.  Figure 1: See a starfish? This is actually a map of Joanne Kamen's LinkedIn... it is completely impossible for me or anyone to stay in touch with all of their connections on a regular...

...different water filtration system, environmental stability in an incubator), but there’s a lot you can do...other researchers to help them troubleshoot and understand. All of this communication and sharing fits together...seriously. That’s why at Cell Press we developed STAR Methods to improve the structure and transparency... of checklists and guidelines to help authors understand what they are expected to do. All of that helps...

...efficiency of these editing reactions is low. Understanding the biological cause of this repair bias is ... reinforce a key scientific principle: that understanding a biological process in detail often suggests... efficiency of sequence replacement by HDR. Understanding Cas9 biophysics enhances HDR We first measured...released strand, 127bp in length, With 36bp on the PAM-distal side of the break and 91bp on the PAM-proximal ...

... the purpose of the meeting: Begin meetings by stating “The purpose of this meeting is to _________." ... what you need to accomplish to get to the next stage, and then adjourn. Define the tone of the meeting...template for project meetings. Team Meetings: For standing team meetings, provide agenda sections for each...meeting. Accountability: At your next meeting, start by checking in on the action items from last meeting...

...Imaging Improving Well-established High-throughput assays Improving Well-established Biological sources...cells. Luciferase assays have since become a gold standard in gene expression analysis and a luciferase gene...Phrixothrix hirtus), and a variety of marine crustaceans. Many of these luciferases have been subjected...

...phenotypes than standard RNAi protocols without the need for mutant genetic backgrounds. The standard RNAi vector...biochemical and biophysical study involving protein stability or binding experiments. Potential experiments ...perform using the purified domains include protein stability, small molecule binding screens, peptide arrays...

...infect cells and deliver cargo, it’s important to understand their differences to choose the best one for ...viruses Retroviruses are a family of viruses that stably integrate their genome into the host genome, setting...   Type of Virus Pros Cons Retrovirus Stable Gene Expression Broad Tropism Only Infects ...virus safety and viral applications to help you start (safely!) thinking about the possibilities. Ready...

...at (and through) a couple of models. Before you start a new project with fluorescent proteins, the best...this series of posts are (i) brightness, (ii) photostability and (iii) aggregation tendency. Brightness...Without further information about properties like photostability or aggregation tendency, our choice would be...Goedhard and Marieke Mastop! Joachim Goedhart is an assistant professor at the Section of Molecular Cytology...

...This method utilizes a packaging cell line that stably expresses Gag-Pol and/or Env, eliminating the need...packaging cell line developed by Garry Nolan at Stanford (Swift, 2001) contains Gag-Pol and either an ecotropic...differentiated (non-dividing) cells. Both lentiviruses and standard γ-retroviruses use the Gag, Pol, and Env genes...provirus. 3’ LTR Terminates transcription started by 5' LTR by the addition of a poly A tract just...