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...sometimes localize to unintended sites with sequence similarity to the on-target site. When a CRISPR-Cas system...frequency of 5%. Your experiment involves making a large sgRNA library to thousands of genes and targeting...targets by selecting a gRNA with low sequence similarity elsewhere in the genome. There are many predictive...Once you’ve selected a gRNA with low sequence similarity elsewhere in the genome to reduce off-targets...to consider your Cas enzyme. Even if there are similar sequences present in the genome, certain Cas enzymes...Candidate site sequencing Sites with high sequence similarity to the gRNA can be sequenced. These sites can...appropriate method for an activator or repressor Cas. Similarly, Cas ChIP-Seq would also identify all DNA binding...

...topics in biology, especially those at the molecular and cellular level. The reason for this is cost and ...it wasn’t until I started working in my first molecular biology lab in college that I really fell in love...limited budget. While there are more and more molecular biology-based activities making their way into... about operating the rest of the car as well. Similarly, cell free expression systems contain all the ...References 1. Huang, Ally, et al. "BioBits™ Explorer: A modular synthetic biology education kit." Science Advances... to learn simple lab techniques Check out our molecular biology reference pages Find plasmids from the...

... by Alycia Bittner CRISPR pooled libraries are popular tools for genetic screens, enabling genome-wide...genome-wide interrogation of the genes required for cellular processes. For a successful screen, it is necessary...Schwiesow A leading cause of death worldwide, cardiovascular disease is driven by atherogenic lipoproteins...lipoproteins that accumulate within the vascular wall. ApoB, the main structural component of atherogenic lipoproteins...been shown to have a strong correlation with cardiovascular disease and indirect measurement of ApoB-LPs...to aid in the understanding and treatment of cardiovascular disease. Find the LipoGlo plasmids at Addgene...78% in body wall muscle cells. These changes were larger than previously reported for other organisms, and...

...used to confirm the presence of an insert in a particular vector by excising the insert from the backbone...insert. You can search NCBI for YGOI to find the particular reference sequence if necessary. The example ...BamHI) are capable of cleaving sequences which are similar, but not identical, to their defined recognition... see 3 bands. This is due to the fact that the circular DNA takes on several conformations the most abundant... the DNA mass of comparably intense samples of similar size. Run the gel at 80-150V until you have good...the gel for longer to achieve good separation of larger DNA fragments. I hope these tips demonstrate ...More Plasmid Eductional Resources: Addgene's Molecular Biology Reference Find More Protocols and Videos...

...including the popular BL21(DE3) (Li et al., 2021). This pEcCas/pEcgRNA system works similar to the original...modifications, efficiency can be low, especially for larger modifications. What’s the solution to this inefficiency...recombinogenic than E. coli, so this system functions more similarly to CRISPR/Cas9 systems adapted for eukaryotes ...promoters are ubiquitous, they are not universal, particularly when it comes to non-model bacteria. The authors...engineer the bee gut microbiome. They created a modular system using a broad host range plasmid that can... specific locus. This approach provides a more granular level of control where individual bases can be...Bee Gut Microbiome Bacteria with a Toolkit for Modular Assembly of Broad-Host-Range Plasmids. ACS Synth...

...face very similar issues when forced to express in mesophilic host systems. The so-called “regular” temperatures...at suboptimal ones, which could allow even your regular E. coli to thrive in the bitter cold.  Since...vs. extremophiles Mesophilic organisms are your regular next-door microbes which grow optimally in moderate...psychrophilic enzymes for milk processing, fruit juice clarification, meat tendering, and oligosaccharide synthesis...the many proteins which are not compatible with regular mesophilic systems are the psychrophilic enzymes...incorporate cold-tolerance inducing genes within a regular mesophilic host system. This would make the host...

...reagents with the scientific community.   (click to enlarge) As we look ahead to the things to come, we wanted...total? Just over 49,000 times! Their long-lasting popularity illustrates the utility these tools have in many... The S. pyogenes Cas9 (SpCas9) has been widely popular in genome engineering. For use in human cell lines...plasmid. Indeed the first version is the 15th most popular plasmid of all time, but was found to have a point...publication but its name was subsequently changed to clarify that it does not contain the trademarked Tet-On...lentiGuide-Puro. pSpCas9n(BB)-2A-GFP (PX461) - Similar to pSpCas9(BB)-2A-Puro (PX458), pSpCas9n(BB)-2A-GFP...gRNA. It’s the former version of our sixth most popular plasmid pSpCas9(BB)-2A-Puro (PX459) V2.0. It contains...

...heavy machinery. But not all engineering is at a large scale — some is done on much smaller equipment on...depth is antibody directed evolution. This is a larger process of gradually changing and selecting the...Degenerate primers Using degenerate primers is similar to error-prone PCR. Degenerate primers are a group...generation. To evaluate improved properties across large mutant libraries, high-throughput screening techniques... Innovation (IPI) provides information on their large-scale recombinant antibody production platform. ...screening, and affinity maturation. Methods in Molecular Biology, 29–62. https://doi.org/10.1007/978-1-...conformationally selective nanobodies. Nature Structural & Molecular Biology, 25(3), 289–296. https://doi.org/10.1038...

...duties Staff scientist: similar role to a postdoc but more permanent Visiting scholar/student: visiting from...conferences. The Barry Goldwater scholarship is also a national scholarship that supports students interested...because they had close relatives suffering from similar diseases. Others were inspired by the imminent ...that you would be interested in can also help you clarify your goals and define next steps! More than serving...online for a while and observed. Apply for those scholarships that you don’t think you’re qualified for. Wayne...populations. Research universities often have scholarships and funds dedicated for undergraduates interested...

...Goedhart and Marieke Mastop from the Section of Molecular Cytology and Van Leeuwenhoek Centre for Advanced... can be excited many times, thereby producing a large number of emitted photons before they are destroyed...absorb the excitation light less efficiently. Similarly, decreasing the power of the excitation light ...these factors, environmental conditions such as cellular redox state and oxygen concentration may affect...Goedhart is an assistant professor at the Section of Molecular Cytology and Van Leeuwenhoek Centre for Advanced...Marieke Mastop is a PhD candidate at the Section of Molecular Cytology (University of Amsterdam) where she develops...Backbones with Fluorescent Tags Find Plasmids for Subcellular Localization Browse the entire Fluorescent Protein...