...Brook Pyhtila One drawback to the commonly used S. pyogenes CRISPR/Cas9 system is that it is too large to ...Find the SEND plasmids    Figure 1: SEND combines an endogenous Gag homolog, cargo RNA, and fusogen...editing to correct mutations in a variety of target genes and cell types. Their Nature Biotechnology paper...

... the same isotype. But together, all the B cell clones in the immune system make different isotypes of... starts with selecting just one of these B cell clones for further antibody production, polyclonal antibodies...antibody increases the likelihood that one of the clones will still recognize your protein. For the same...

...overcome by screening a higher number of potential clones. Golden Gate assembly has a few advantages over... allows for multiple DNA components (promoters, genes, terminators, etc) to be assembled in the correct...ligated following a heat inactivation step. If your genes of interest or destination vector contain multiple...

...malignant cell, then you’d need the ability to turn on genes at different times. It requires more complex synthetic...on an engineering project including hundreds of genes and regulation and resource control - everything...gives us enough hardware to control hundreds of genes. We’ve created enough regulatory elements, CRISPRs...

...Nme) (Pawluk et al., 2016). While Streptococcus pyogenes Cas9 (SpyCas9) is the most used and well studied... Selection marker while engineering viruses Acr genes can be used as positive selectable markers in engineering...viruses. For example, an Acr gene was used to replace genes in the difficult-to-engineer Sulfolobus islandicus...

...with those particles. Since the Env, Gag, and Pol genes are not encoded in the transfer plasmid, the component...also have a propensity to integrate into proto-oncogenes, which can lead to cancer. These viruses need ...attention to the trainings!), even if you know guidelines for best practices and have been trained elsewhere...

...achieved using lentiviral backbones expressing blasticidin or puromycin resistance genes.  It’s that simple...Vogelstein B (1983) Hypomethylation distinguishes genes of some human cancers from their normal counterparts...

...institute’s blog. The Wellcome Trust blog or the John Innes Student Voice are two examples. Many blogs like ...Londonist. BBC provides a peak into their day-to-day business with popular work experiences of 3-10 days. And...started writing three years ago on a field trip in Indonesia. Since then, I have tried many things: photography...

...Plasmids designed to express genes in a given host cell type are generally broken down into two broad...Increases Expression Levels of Lentivirus-Delivered Transgenes but Has the Potential to Reduce Viral Titer in...Posttranscriptional Regulatory Element Enhances Expression of Transgenes  Delivered by Retroviral Vectors. Journal of ...

...management-for-scientists-managing-vs-leading. The National Postdoctoral Association (NPA) outlines 6 Core Competencies that all postdocs (but also...actionable tasks and I made a point of sticking to deadlines and performing the work I agreed upon (just as...accommodate many different types of people in order to harness their strengths and work effectively. Leadership...

...panelists will negatively affect timing and the liveliness of the discussion (see below on timing). Get ...what happens to the attention of the audience – phones come out, postures shift, they are checking out...more great tips on running panel discussions see Guidelines for Great Panels by Scott Kirsner and How to ...

...system, in which the required adenoviral helper genes are supplied on two different packaging plasmids...pHelper plasmid containing Ad E2A, E4, and VA RNA genes; and the pRepCap plasmid containing replication ...replication factors and serotype-specific capsid genes.  In this helper-free system, HEK293T cells, expressing Ad...

...The degradation tag (dTAG) system functions by harnessing the power of a novel degrader of the 12 kDa protein...round of evolution (AID-2) to reduce some of the leakiness associated with the system, but the general concept...all the tags discussed here and profile their effectiveness with over a dozen different proteins. In the...

...Manuel Leonetti to tag all 22,000 protein coding genes in the human genome with fluorescent proteins. For...tool Crispor by Max Haeussler. With a few extra lines of code, we enabled it to return guides and primers...which is important for quality control. The only ones we found that worked with NGS data were: Crispresso...

...reaction but can be used to quickly generate multiple clones. Difficulty: If you’re an experienced cloner, you...also quite efficient; although efficiency often declines as the number of fragments you’re hoping to assemble...out protocols for plasmid cloning Find vector backbones Read our molecular biology reference ...

...and departments will have qPCR machines available for use. ddPCR machines are not as commonly available...measurement of the low-affinity Fc gamma receptor genes FCGR3A and FCGR3B. PloS one, 10(1), e0116791. https...

...generating mutated sequences for site-directed mutagenesis.  Quantitative polymerase chain reaction, qPCR...interest  Molecular Cloning Site-directed mutagenesis Molecular Diagnostics qPCR Quantify starting... PCR  Plasmids 101: Colony PCR Site Directed Mutagenesis by PCR Resources on Addgene.org Protocol: Polymerase...

...are millimeters in thickness (8,9).  In addition, there is a drop in image brightness with increasing depth... Ultramicroscope.  In addition, adjusting the thickness of the laser sheet and using multi-view imaging...

...should be run in duplicate or triplicate. The closeness of the data points generated by replicate values...be linear or nonlinear. BCA assays have linear trendlines while those for ELISAs and other enzymatic assays...advanced graphing software designed to plot complex trendlines. If you don’t have access to this kind of software...

...editing from the Cas13b REPAIR system To test the robustness of REPAIRv1, the lab targeted 34 pathogenic G...cytosine deaminase. Three rounds of rational mutagenesis on the ADAR2DD residues that contact the RNA ...RNA-sequencing, the authors performed rational mutagenesis and identified that a S375A mutation in RESCUE...