...editing!  A recent paper from Feng Zhang's lab describes how to use Cpf1 for multiplex genome editing.  ...transfection gRNAs expressed from a polycistronic transcript  Requires co-transfection of Cas9 Nissim et alTsai...transfection gRNAs expressed from a polycistronic transcript Requires co-transfection of Cas9   Xie et al  ...

...expressing cell lines, which can then be used both for screens (using pooled CRISPR knockout libraries) and individual...monoclonal Cas9-expression line prior to performing a screen or even a single gene knockout. Top AAV: pAAV-hSyn-DIO-hM4D... vectors and genome-wide libraries for CRISPR screening. Nat Methods. 2014 Aug;11(8):783-4. PubMed PMID...

...Cell, Musunuru, Chad Cowan and their colleagues describe a much more efficient tool for doing that disease...modeling work in human pluripotent stem cells: a transcription activator-like effector nuclease (TALEN) kit...people unfamiliar with this new TALEN kit, can you describe it for us? Musunuru: TALENs are a type of genome...

...formulation of new knowledge than it is to read a manuscript whose whole purpose is to provide detailed evidence...images, noises and music, spoken words, and any onscreen text. Be aware of the effort the audience is expending...Maybe I needed to employ a visual aid instead of describing something verbally. Sometimes a group was just...

...Systems Biology in November, Voigt and Alex Nielsen describe what’s possible when multi-input CRISPR/Cas genetic...many deposits to Addgene over the years. Can you describe the toolbox you are putting together?  Voigt: ... Those will allow you to set the amount of transcriptional and translational activity a cell has and control...

...soCoChR-expressing system. Taken together, Shemesh et. al. describe a novel system for the precise mapping of neuronal...the single-cell level. Various soCoChR plasmids described in this work including the soCoChR-GFP plasmid...expression in E. coli. This operon is under the transcriptional control of a strong constitutive promoter (...

...neurons from each class to fully characterize the transcriptome. The first data release for CeNGEN is planned... The 3’UTRs of mRNA are important for post-transcriptional regulation in C. elegans. To study the role...Addgene In addition to the wealth of open resources described above, there were many plasmid tools presented...

...'ve overlooked regulatory fragments of these transcripts. I remember when small RNA was just something...’t give tRNAs much thought. High throughput transcriptome sequencing methods including work done by the...circRNA datasets and download the custom python scripts needed to discover circRNAs in RNA-seq data. As...

...histone modifications, which can influence the transcription of genes by RNA polymerase II, or by the addition...methylcytosine in CpG islands makes  genes less transcriptionally active. The role of epigenetics in studying...of tumor suppressor genes can repress their transcription and inactivate them. Global hypomethylation,...

...synthesize a particular protein DNA must first be transcribed into messenger RNA (mRNA). mRNA can then be translated...template of choice and allowing these cells to transcribe, translate and modify your protein of interest...assembled in vitro using purified components of the transcription and translation machinery. These include ribosomes...

...sequences that, when bound by transcription factors, can increase a gene’s transcription. Sometimes enhancers ...vivid chromatin picture, CAPTURE 2.0 re-writes the script on gene regulation. References Han, J., Zhang...

...chromoproteins have a shorter history. In 1987, scientists described an unusual blue-colored protein from the jellyfish...methods. While techniques such as  blue-white screening require X-gal, an expensive exogenously-added ...chromoproteins don’t require other substrates for screening. Thus chromoproteins can be used as markers in...

...transduced cells whenever possible. This post will describe our workflow for viral genome sequencing (VGS)...identification, we developed custom open-access Python scripts that can perform a deeper analysis of the data....Check out our Viral Vectors Featured Topic Page Subscribe to Viral Vectors Blog Posts Learn How We Use NGS...

...transfer plasmid at 30 °C instead of 37 °C and then screen for ITR recombination with a SmaI or XmaI restriction...duplicated regions like the ITRs. Restriction digest screening for ITR deletion is still necessary. To save time...mL of my AAV transfer plasmid cultures and then screen with SmaI digests. I freeze the bacterial pellet...

...following cloning methodologies have been extensively described in other articles; so if you are interested in... to assemble up to 6 different fragments in a prescribed order. These fragments can be pooled in one tube...homology. You might also consider FastCloning, first described by Chaokun Li and colleagues. Here you simply ...

...vivo. A common choice for conducting genome-wide screens using CRISPR. AAV transduction Only compatible...containing gRNA and Cas9 are used in in vitro transcription reactions to generate mature Cas9 mRNA and gRNA...complexes Purified Cas9 protein and in vitro transcribed gRNA are combined to form a Cas9-gRNA complex...

...looking at your resume while they wrote the job description. You check off all the boxes - you have the desired... gaps in scientists' transferrable skills. In describing the limitations of the study, the authors noted...over email. Examine your current and past job descriptions. Scan through all the things for which you are...

...video_player "embed_player" overrideable=False, type='scriptV4', hide_playlist=True, viral_sharing=False, embed_button...into two components: 𝜃x(t) and 𝜓z(t). 𝜃x(t) describes the exponential decay process we observed, while...appears to return to normal. The model fit these two discrete phases with an estimate of 62.3% for the COVID...

...amplification) or reverse transcriptase (RT)-RPA, respectively. RPA is coupled with T7 transcription to convert amplified...2018) CRISPR-Cas12a target binding unleashes indiscriminate single-stranded DNase activity. Science 360...

...we probably exchanged dozens of drafts of the manuscript. This is the time to transform the document into...Resources on the Addgene Blog Writing Scientific Manuscripts: Literature Searching, Reading, & Organizing ...Organizing Tips for Improving Your Next Manuscript 6 Tips for Grant Writing Resources on Addgene.org Check out...