...and easily generate zebrafish transgenic lines to study neurobiology and neuorodgenerative diseases. The...Gateway-Compatible Toolbox is based on the original Tol2kit generated by the Chi-Bin Chien lab (Kwan et al., 2007) and... specific neuronal cell types involved in neurodegenerative diseases, the Cole Lab selected promoters ...The impact of KLF4 N-terminal variants on iPSC generation Deriving iPSCs (induced pluripotent stem cells...reprogramming somatic cells. A common method for generating iPSCs to express the transcription factors from...Polycistronic cassettes that contained KLF4s had overall diminished KLF4 expression levels; this altered...mScarlet. Bindels et al used a combination of deliberate and random mutagenesis to isolate the best RFP...

...Promoters control the binding of RNA polymerase and transcription factors. Since the promoter region ...protein, the repressor protein can bind to the operator. The bound repressor then prevents transcription... repressor (TetR) can bind to the tetracycline operator sequences (TetO), preventing transcription. In...have been engineered and improved. Takara’s “2nd generation Tet-Off vectors (Tet-Advanced)” are the most ...VP16. cTA binds to promoters containing putative operator sequences (CuO) (linked in groups of 6), allowing...the same manner, with LexA binding to the lexA operator (lexAop). Fusing the LexA DNA-binding domain (...refined the Q system and introduced the second generation QF2 activators. These activators solve the problem...

...piggyBac™ transposase or hyperactive transposase mutants [7], it is generally not as efficient as Cre ...How does the piggyBac™ system make it easier to generate transgenic cell lines? One of the major uses of... that have negative impacts on fitness. One consideration when making a transgenic cell line with the ...nucleases (CRISPR/Cas9, TALENs, and ZFNs). However, several challenges still remain, such as making gene edits...completely seamless (i.e., so no other genetic alterations have occurred) and isolating the specific cells...the piggyBac™ system, these challenges can be considerably reduced. The piggyBac™ transposon system can...and M. H. Wilson, “piggyBac-ing models and new therapeutic strategies.,” Trends Biotechnol., vol. 33, no...

...ion-channels and pumps in developmental biology, regeneration studies, and cancer biology. I was first introduced...patterning. Based upon trends in studies over the past several years, it seems that the more polarized (negative...  Additionally, researchers have reprogrammed regenerating tissues without using genome engineering. Planarian... briefly inhibited in the entire worm during regeneration, the worm creates two heads. Subsequent cutting...transcriptional or translational level. However, several genetic tools have been developed to address these...exciting emerging field, with many implications for regenerative medicine, evolution of body plans, synthetic...the advancement of research in birth defects, regeneration, medicine, and synthetic bioengineering. References...

...produced from its serum several weeks later. Created with biorender.com. To generate polyclonal antibodies... response.   Polyclonal antibodies are generated by injecting an animal with an immunogen, then...might be the full-length protein that you want to generate antibodies against, or a unique peptide sequence...Freund’s adjuvant is injected, which stimulates the overall immune response. Once the initial antibody response...designed for experiments where antibodies raised in several different species are present. One common example...antibody for your experiment, you should either order several vials of the same lot of polyclonal antibody, or...experiments to be aware of signal strength. In general, polyclonal antibodies produce a higher level of...

...(tracrRNA) which interacts with the Cas9 protein. These RNA molecules are vulnerable to attack by both...effectively serving their function of guiding Cas9 to generate a targeted break. These advances in gRNA technology...foreign RNA. To protect the RNA from degradation, several simple methods have been employed which modify ...modification is best? Studies suggest coupling several of them together, for example phosphorothioates...more efficient on a budget is to simply swap out several of the ribonucleotides for deoxynucleotides. A ...A partially DNA gRNA is surprisingly well-tolerated by the Cas9 protein and can increase on-target efficiency...photocleavable gRNAs. Photocleavable guides are generated by introduction of a single photocleavable 2-nitrobenzyl...

... islandicus CRISPR-Cas system. 5. Phage therapy Phage therapy uses phage to treat bacterial infections...have discovered over 50 anti-CRISPR proteins that interact with CRISPR-Cas systems such as Cascade-Cas3, ... DNA binding Example:  AcrIIA4 blocks Cas9’s interaction with the PAM site (Dong et al., 2017), and AcrIIC3...2017). Inhibit crRNA loadingExample: AcrIIC2 interactions with Cas9 interferes and prevents the correct...bacterial pathogens possess CRISPR-Cas systems, phage therapy might not be as effective. This is where Acr proteins...enough that they could be engineered to “armor” therapeutic phages to protect them from pathogenic bacteria...activity is an important step towards CRISPR-based therapies and for more precise editing in the lab. References...

... top of the ever expanding body of biomedical literature! PubMed PubMed is the classic search engine ...Twitter into a paper-finding-machine. There are four general types of accounts you can follow: Scientists: ...updated. This Twitter list contains >300 Twitter literature bots, including a bot for biosensor papers, microbe...Semantic Scholar Semantic Scholar is an AI-backed literature search developed by the Allen Institute for Artificial...search terms. Key features Semantic Scholar has several ways to focus your search results. A bar graph ...quickly identify papers with greater influence, the average number of citations per year and the number of ...aren’t familiar with the topic.  Meta Meta is a literature discovery platform developed by the Chan Zuckerberg...

... with single-pot assembly Tracking cell-cell interactions New auxin inducible degron system for C. elegans...    New system developed to track cell-cell interactions By Jennifer Tsang GFP-based Touching Nexus (...Winslow’s lab to label cells after cell-cell interactions. Using G-baToN, “sender” cells express GFP on...anti-GFP nanobody on the cell surface. When the cells interact, GFP is transferred to the receiver cell, and ...cell pairs.   Figure 2: Schematic of the interaction between the sender cell and the receiver cell...with BFP, sender cells expressing surface BFP generated BFP-labeled receiver cells. What’s more, G-baToN...provides versibility in tracking and modulating the interactions between cells.      Tang et al., eLife 2020...

... to consider one of several Cas9s that recognize NG PAMs. The Nureki lab generated SpCas9-NG, a rationally...biology, the answer is always it depends, but generally the efficiency of the desired edit is reduced ...name implies (Nishimasu et. al, 2018). xCas9 was generated by the Liu lab through phage-assisted evolution...evolution, specifically with the goal of generating Cas enzymes with alternative PAM requirements (Zhao et. ...through structure-guided engineering of the PAM-interacting domain of Cas9. After multiple rounds of engineering...helps even when it’s not an ideal site: moving several bps away from a desired site does not have the ...most gene targeting experiments. Between SpRY and several of the other Cas variants described, nearly all...