...in more genome engineering technologies? Browse others on Addgene's Genome Engineering Guide. Fire ...gene targeting Victor de Lorenzo's lab has engineered a modular mini-Tn5 vector that can be used to...biochemical characterization, has enabled scientists to engineer a wide variety of GFPs with diversity in fluorescence...transcriptomes, as well as further optimization through engineering, Ed Boyden’s lab has identified a variety of ...implicated in disease progression. One of the major challenges in signaling research is the inability to replicate...field of optogenetics. Using a rational protein engineering approach, they designed Opto-RTKs, which activate... has now made improvements to this system by engineering cells that lack release factor one (RF-1; Bacterial...

...conditions. Shyu YJ, Liu H, Deng X, Hu CD. Biotechniques. 2006 Chang-Deng Hu Green sfGFP Split super-folder...conditions. Shyu YJ, Liu H, Deng X, Hu CD. Biotechniques. 2006 Chang-Deng Hu Venus Improved N-terminal...Split-Cas9 for inducible genome editing (Feng Zhang Lab 2015)  Reference Feng, S., Sekine, S., Pessino, V. et...producing fluorescent signal. Image modified from Feng et al. 2017 under CC-BY license. Many split-FP...fluorescent proteins for endogenous protein labeling. Feng et al. Nat Commun. 2017 Bo Huang mNeonGreen3 mNG3K...protein labeling and signal amplification. Zhou S, Feng S, Brown D, Huang B. PLoS One. 2020 Bo Huang Yellow...ratio. Kodama Y, Hu CD. Biotechniques. 2010 Chang-Deng Hu mVenus pET-BiFC contains both fragments of ...

...: A systemic capsid is a capsid that has been engineered to have enhanced function after intravenous administration... post, we’ll discuss how systemic capsids are engineered, as well as highlight some considerations for...region-specific transgene expression. (a) Several engineered capsids with unique tropisms are now available... rodents and non-human primates (NHPs). These engineered capsids have increased transduction efficiency...generally compared to the AAV9 serotype or an engineered precursor of the systemic capsid in question ...better efficacy than AAV9 and/or its intermediate engineered precursor. Considering systemic capsids for your...determined by the capsid and promoter/enhancer strengths in the cell type of interest as well as the type...

...light-pulse at one wavelength and stay on until a second pulse of light, with a different wavelength, turns it ... for your experiments.  Wavelengths Most vectors have an optimal wavelength to use but can be activated...of other cellular functions. Photoswitches are engineered molecules that induce conformational changes ...exposure to light within a specific range of wavelengths. Excitatory channels allow positively charged...and inhibitory channels activated at different wavelengths of light (Vierock et al., 2021). Step-function.... There are many different opsin (natural and engineered proteins responsive to light in combination with...designed to be activated within a narrower band of wavelengths to allow multiple tools, such as biosensors, ...

...and then derive mice from these engineered ESCs. To do so, the engineered ESCs would be injected into developing... the cells in the adult mice derived from the engineered ESCs), and chimeric adults mated to produce completely...Mouse Modeling blog posts: Part 1: Genetically Engineered Mice and Part 2: Breeding and Crossing Mice. ... to CRISPR when he used this new technology to engineer mouse models (Wang et al., 2013, Yang et al., ... Advantages of using CRISPR for mouse genome engineering From the very start, generating mouse models ..., compared to conventional methods of genome engineering, CRISPR is much more efficient at generating ...Marraffini LA, Zhang F (2013) Multiplex Genome Engineering Using CRISPR/Cas Systems. Science 339:819–823...

... specific wavelengths of light to initiate signaling. When FRET donor excitation wavelengths overlap with...proteins in whole animals, light at or above 600 nm wavelengths should be used. Researchers often use bioluminescence...traditional luciferases emit at near optimal wavelengths (Fluc emits at 610 nm) and do not require external...fluor then emits light at a tissue-penetrating wavelength above 600 nm. LumiFluors Schaub, F.X., et al....that could be co-imaged with GFP using the same wavelength for excitation. The result was the cyan-excitable...This strategy was first applied using RLuc8, an engineered Renilla luciferase, to produce a series of FP-RLuc8...concentrations of biological molecules. FRET sensors face challenges of photobleaching, autofluorescence, and, in ...

...optogenetics.  Challenges of engineering channelrhodopsins There are three main challenges to engineering channelrhodopsins...overcome channelrhodopsin design challenges To overcome these challenges, the Arnold and Gradinaru Labs ...predict the channelrhodopsin’s photocurrent strength, wavelength sensitivity, and off-kinetics.   This model...channelrhodopsins with high-throughput proteins engineering methods, like directed evolution, because channelrhodopsins...functional and their measured photocurrent amplitude, wavelength sensitivity, and off-kinetics correlated with...lower multiplicity of infection (MOI) makes it challenging for ChR2(H134R) to function because it’s lower...2019) Machine learning-guided channelrhodopsin engineering enables minimally invasive optogenetics. Nat ...

...PMC6342934. Teng, Fei, et al. "Repurposing CRISPR-Cas12b for mammalian genome engineering." Cell discovery...of Cas9s isolated from different organisms and engineered Cas9 variants that were able to target different..., the Osamu Nureki Lab described a rationally engineered Cas9, SpCas9-NG, which required only the di-nucleotide...time of our first blog post on choosing a Cas9, the Feng Zhang Lab had described eSpCas9, while the Keith...different Cas9 variants using gRNAs of different lengths (Lee et al., 2018) and found that Sniper-Cas9 showed...sensitivity across a large number of loci and gRNA lengths. Recent alternatives to Cas9 While Cas9 remains...characterized by the Zhang (Strecker et al., 2019) and Li (Teng et al., 2018) Labs also shows promise for in vivo...

... you generate a growing colony of genetically engineered mice, what can you do? In this post, we’ll cover..., and how to properly control for genetically engineered mice in your experiment. As you’ve learned in...in Part 1, there are many types of genetically engineered mice: transgenic mice, knockin and knockout mice...generate a desired phenotype Instead, genetic engineering should be used to separately introduce each mutation...offspring that have a combination of the mutations engineered into each of their parents. Here’s how to cross...Whether your mouse colony consists of genetically engineered mice with a single mutation or strains that have...can be mitigated in different ways. Prior to engineering a transgenic strain, the promoter being used ...

...characterizing the strength a species absorbs or reflects light or radiation at a specific wavelength, and the ...light or electromagnetic radiation at another wavelength (Heinrichs, 2009). When a fluorophore absorbs...difference in emission results in a slightly longer wavelength emitted than was absorbed. This phenomenon is...known as the Stokes Shift, or the difference in wavelength between peak excitation spectrum and emission...differences in their excitation and emission wavelengths, which are pivotal in their application in biotechnology...Hayashi-Takanaka et al., 2014). Other dyes may prove challenging to stain intracellular compartments in in vivo...applications that involve ranges of pH, and significant lengths of time, the stability of fluorescence over time...

...Cas family of proteins has many useful genome engineering enzymes, each with their own unique features ... and quirks. Here, we will review the genome engineering abilities of the lesser-known Cas enzymes while... DNA targeting Cas enzymes Cas9 – The genome engineering workhorse You rarely hear the word ‘CRISPR’ without...hearing ‘Cas9’. The Cas9 nuclease is a genome engineering powerhouse, and for good reason. It is guided...strand break. Cas9 has been studied for genome engineering purposes far more than any other Cas family protein...making Cas9 an adaptable choice for many genome engineering endeavors. For more information on Cas9 variants...curious feature of this Cas is that once it is engaged by its target, it will non-specifically cut ssDNA...

...reprogramming  Regulation of telomere length with TZAP Maintaining telomere length is important for cellular function...on regulating telomere length with TZAP Multiplexible plant genome engineering toolkit The Voytas Lab ... Segment on PhoCl LSSmCherry1 & RDSmCherry1: Engineering and directed evolution of new RFP variants The...were developed as part of a larger strategy to engineer improved red fluorescent proteins (RFPs) and, ... of base editing. Kim et al. used natural and engineered Cas9 variants to develop five new base editors...The Denchi lab investigated an aspect of telomere length regulation called telomere trimming and have proposed...ZBTB48) in regulating the upper-bound of telomere length. Telomere trimming can be assayed by looking for...

...has been a lot of engagement with the Athena SWAN Charter and it has really strengthened the network of ...Laboratory, where she often feels more like an engineer than a biologist. Their focus at the lab is on...overcome them, and why she spends some of her time engaging with others on Twitter, among other things. MoClo...fluorescent proteins. Visualizing fluorescence can be challenging in plants because of the auto-fluorescence of...similar resource for those involved in monocot engineering projects. The MoClo tools will work in any dicot...science, and many other topics. What’s the value of engaging about science in that way for you? Patron: I started...Gate Modular Cloning Toolbox for Plants. Carola Engler, Mark Youles, Ramona Gruetzner, Tim-Martin Ehnert...

...The emitted light is at a longer wavelength than the exciting wavelength. Thus, FPs are proteins with this...by a specific excitation wavelength. This means that the emission wavelength can change. In a few cases...FPs classified by the emission color (emission wavelength range) FPs are usually classified by emission...emission color as outlined below (or emission wavelenght range).  By mutating GFP, the variants blue FP (BFP...Infra-Red ≥ 670 nm In addition to emission wavelength range, there are other traits that need to be...named after George G. Stokes) is the shift in wavelength from excitation to emission. For most FPs, Stokes...sure their emission light does not overlap in wavelength. In many microscopes the filters are not narrow...

...Open Yeast Engineering Project (OYEP). Through this project, community members will engineer yeast to glow...that anyone can engineer yeast to do similar things. The focus is to make yeast engineering accessible to...are skilled with audiovisual media, software engineering, business consulting, mathematics, and more. ...and in his free time, he is pursuing a plant engineering project at BosLab. Once he optimizes plant cell...knowledge with the community by teaching a DIY plant engineering workshop. Community projects at BosLab Not all...became a member at BosLab because she wants to engineer microbes. We’ve ordered a few yeast plasmids from...diversity in different soil conditions/locations but to engage the local community (including local garden and...

...BioBuilderClub BioBuilderClub engages high school teams around the world who combine engineering approaches and scientific...are asking. Teachers are also asking, “How do I engage my students in real-world lab activities? I’m trying... Kuldell from MIT’s Department of Biological Engineering. Built primarily from her 20 years of college...classroom are characterized by intense student engagement and joy, rather than a perfect lesson plan or... boost interest and understanding around the engineering of biology. Participating schools are provided...BioBuilder inspires students and teachers to apply engineering principles to biological systems. Today, what...BioBuilder module allows students to explore how engineering fields rely on rapid prototyping to develop and...

...the Wyss Institute for Biologically Inspired Engineering. CRISPR/Cas9 technology has revolutionized the...the fields of molecular biology and bioengineering, as it has facilitated the development of a simple ...different perturbations. Consider, for instance, the challenge of attempting to simultaneously and selectively...of modulating gene expression (6). Altering the length of gRNAs targeting different sites of interest ...orthogonal Cas9 species. Within a single cell, full-length guides targeting one set of genetic loci can be...Santa Cruz, where her research is focused on engineering nanoparticles as drug delivery systems. She is..., Raj, et al. "Unraveling CRISPR-Cas9 genome engineering parameters via a library-on-library approach....

...nuclease domain. Feng Zhang's group utilized the bi-lobed architecture of Cas9 to engineer a series of "split...memory One of the goals of synthetic biology is to engineer cells capable of recording permanent "memories...display and high throughput binding assays, to engineer photoswitchable dimers which show over a 50 fold...activating endogenous genes Moritoshi Sato's Lab has engineered CRISPR-Cas9 to create a photoactivatable transcription... or (4) multiple sgRNAs with Cas9 for genome engineering. The versatility and ease of generating complex...potential applications of CRISPR/Cas9 in genome-engineering. One particularly attractive application involves...genes involved in neurogenesis, which had proven challenging using the existing dCas9-VP64 activator. The ...

...Addgene to advance their research. Louis Bengyella Dr. Bengyella’s work has focused on elucidating the molecular...our first two plasmid grant awardees: Drs Louis Bengyella and Kwabena O. Duedu, both from the University...in Ghana. Read on to learn more about doctors Bengyella and Duedu and how they plan to use plasmids from... microbial pathogens. This work has given Dr. Bengwella expertise in using bioinformatic and proteomic...associated with pathogenesis. In his future work. Dr. Bengyella would like to use the expertise he’s developed...development of uterine tumors. According to Dr. Bengyella, this class of tumors is “a major public health... Ghana. We’re excited to see what impact Dr. Bengyella’s work will have on women’s health in Ghana and...

...funding, bioethics in the age of CRISPR, and the challenges of science becoming increasingly politicized...., “The Professional Scientist,” addresses the challenges of succeeding in science if you’re anything other... course the rapidly changing world of genome engineering via CRISPR.  Though the individual topics are...ramifications. Readers unfamiliar with academia’s challenges with publication and funding may be dismayed ...The three chapters in this section describe the challenges of trusting science in a “fake news and post-...has played in these fields, and seeing names like Feng Zhang, Jennifer Doudna, Karl Deisseroth, and Ed ...describes the author’s thoughts on science’s biggest challenges over the coming years. It is arguably the most...