... Protocols Inoculating a Liquid Bacterial Culture Inoculating a Liquid... for Inoculating a Bacterial Culture Protocols...experimental use. The following protocol is for inoculating an overnight culture of liquid LB with bacteria...Watch the protocol video below to learn how to inoculate bacteria in liquid culture. Protocol Prepare liquid...is LB media + antibiotic without any bacteria inoculated. You should see no growth in this culture after...µg/mL Gentamycin 10 µg/mL Kanamycin 50 µg/mL Spectinomycin 50 µg/mL Tetracycline 10 µg/mL Tips and FAQ ...

...mouse and human kinomes. While any subset of the genome is limited by definition, the kinome is a well-annotated... About CRISPR Pooled Library Screening Why the kinome? While properly designed perturbations – in the...incremental approach is prudent. What's in the CRISPR kinome library? Thus, we are glad to provide CRISPR libraries...individual lentiGuide gRNA plasmids targeting the human kinome. These plasmids contain the same 3-4 gRNAs found... kinome...

...Image adapted from Ino et al. 2022, under CC-BY license.   Find MTRIA sensors here!   Ino D, et al. A fluorescent...develop a biosensor for oxytocin, Daisuke Ino and Hiroshi Hibino and colleagues devised a “Modular fluorescence...G-protein-coupled receptor (GPCR) recognizing a specific ligand (Ino et al. 2022). The oxytocin sensor MTRIAOT enabled...

...redirecting specific codons to encode amino acids beyond the 20 standard amino acids. Expanding the Genetic Code...translation, an aminoacyl-tRNA synthetase (aaRS) loads its cognate tRNA with a specific amino acid. Then, ...cognate aminoacyl-tRNA-synthetase (aaRS) that has been evolved to incorporate non-canonical amino acids ...incorporate a non-canonical amino acid into the protein of interest: The non-canonical amino acid, which is generally... recognizes this codon. An aminoacyl-tRNA synthetase to load the new amino acid onto the tRNA. The tRNA...pANAP AnapRS E. coli 3-(6-acetylnaphthalen-2-ylamino)-2-amino-propanoic acid (Anap) Mammalian TAG Peter ...structurally analogous amino acids Bacterial TAG Ryan Mehl 85502 pDule-para-aminoPhe pAF synthetase M. jannaschii...

...PRKCG MTH Spinocerebellar ataxia 30 Frederic Mushinski 8418 pLTR PKC gamma PRKCG Spinocerebellar ataxia ...GFP CMV Spinocerebellar ataxia 27 Tobias Meyer 21204 GFP-N2-PKCgamma PRKCG GFP CMV Spinocerebellar ataxia...GFP CMV Spinocerebellar ataxia 25 Tobias Meyer 21215 GFP-C2gamma PRKCG GFP CMV Spinocerebellar ataxia ...CMV Spinocerebellar ataxia 23 Bernard Weinstein 21237 PKC gamma dNPS PRKCG HA CMV Spinocerebellar ataxia... CMV Spinocerebellar ataxia 21 Bernard Weinstein 21239 PKC gamma DN PRKCG HA CMV Spinocerebellar ataxia...CMV Spinocerebellar ataxia 15 Ryohei Yasuda 112270 CMV-PKCg-mEGFP-N1 PRKCG GFP CMV Spinocerebellar ataxia...Multisystem proteinopathy, ALS Nicolas Fawzi 139121 hnRNPA2_LC_NtoS HNRNPA2B1 T7 Multisystem proteinopathy, ALS...

...ULBP4, bA350J20.7 RAET1G retinoic acid early transcript 1G ULBP5 RAET1L retinoic acid early transcript 1L... PIK3CD phosphoinositide-3-kinase, catalytic, delta polypeptide p110D PIK3CG phosphoinositide-3-kinase...PF4A, SCYB4V1 PLAU plasminogen activator, urokinase ATF, UPA, URK, u-PA PLAUR plasminogen activator, urokinase...KGFLP1 keratinocyte growth factor-like protein 1 MGC125746, MGC125747, MGC126891 KGFLP2 keratinocyte growth... DG6, PMBP PLAU plasminogen activator, urokinase ATF, UPA, URK, u-PA PLAUR plasminogen activator, urokinase...protein 2 FLJ23282, FRA RARA retinoic acid receptor, alpha NR1B1, RAR RARB retinoic acid receptor, beta HAP...NR2B1 RXRB retinoid X receptor, beta DAUDI6, H-2RIIBP, MGC1831, NR2B2, RCoR-1 RXRG retinoid X receptor...

...attached to Venus via a 5 amino acid linker C17V Cerulean attached to Venus via a 17 amino acid linker C32V Cerulean...attached to Venus via a 32 amino acid linker C5A Cerulean attached to Amber via a 5 amino acid linker C17A Cerulean... to Amber via a 17 amino acid linker C32A Cerulean attached to Amber via a 32 amino acid linker mGFP-10...Monomeric EGFP attached to super-REACh via a 10 amino acid linker ACA Heterotrimeric construct consisting...two Venus fluorescent proteins connected via a 5 amino acid linker V17V Anisotropy/brightness standard ...two Venus fluorescent proteins connected via a 17 amino acid linker V32V Anisotropy/brightness standard ...two Venus fluorescent proteins connected via a 32 amino acid linker VVV Anisotropy/brightness standard consisting...

...resource to have in your pocket.   5. +AMINO- by iGreenEarth AMINO is a very simple app that packs important...abbreviations of the amino acids memorized, but the structures are another story. AMINO has been very helpful...properties of or draw an amino acid or peptide for some reason. It contains all twenty amino acids plus selenocysteine...wishing that it had a codon list for each amino acid, but, overall AMINO can be beneficial for new researchers...important biological information, amino acid abbreviations, and structures into one convenient resource. ... Promega Material Safety Data Sheet by RSS-ICTD Amino Acid Structure Quiz/Card by Andrey Solovyev Lab ... He works with gene expression in Pseudomonas aeruginosa. Follow him on twitter @100percentSDS.     Additional...

...in which non-canonical amino acids (ncAAs, also sometimes called unnatural amino acids) are placed in a...re probably quite interested in the canonical 20 amino acids. But in your quest to learn more about your...protein of interest, you may find the available amino acids a bit…limiting. It may be time, then, to look...allowed for intentional incorporation of an unnatural amino acid in translation (Noren et al., 1989). In the...developed into an E. coli system, with the noncanonical amino acid in the media and the tRNAs and synthetase introduced...First, and most obviously, you need a non-canonical amino acid, which is usually added to the media for uptake...result in the incorporation of the non-canonical amino acid at the site of interest (figure 1). While the...

...genes encoding amino acid biosynthetic enzymes may be preferentially charged during amino acid starvation...preferences for the codons they use to encode specific amino acids. This is possible because there are 4 bases...There are therefore 64 possible codons but only 20 amino acids and 3 stop codons to encode leaving 41 codons...result is redundancy; multiple codons encode single amino acids. Evolutionary constraints have molded which... codon tables showing which codons encode which amino acids (see example to the right). With such simple...correctly, and charge the tRNAs with the appropriate amino acids. If an organism uses only a subset of codons...starvation thus resulting in higher production of amino acid biosynthetic enzymes (Dittmar et al., 2005). ...

...of as little as 8 or as many as several hundred amino acids, depending on the tag, and are introduced ...and cons. FLAG FLAG is a synthetically derived 8 amino acid tag (DYKDDDDK) that can be added multiple times...protein with enterokinase, which recognizes the 5 amino acids on the C terminal of the tag. Of the most ...with 5 negatively charged and 2 positively charged amino acids. FLAG tag has been used with moderate to high...as a chimera (rabbit). Myc The Myc tag is a 10 amino acid peptide (EQKLISEEDL) derived from the protooncogene...with 4 negatively charged and 1 positively charged amino acids. Myc tags are frequently used for western ...for all of these species. HA The HA tag is a 9 amino acid peptide (YPYDVPDYA) derived from human influenza...

...G418. Con: Slow transformation   Spectinomycin/streptomycin Spectinomycin and streptomycin both inhibit ...selection agent. Pros: Stable (spectinomycin). Cons: Cost (spectinomycin) or stability (streptomycin), ...Expensive.   Kanamycin Kanamycin, or kan, is an aminoglycoside which inhibits protein synthesis. The kan resistance...The NPTII gene confers resistance to several aminoglycosides, including G418 which is commonly used for ...carb (they are interchangeable). Like carb, spectinomycin is the more stable and expensive antibiotic ...antibiotic compared to streptomycin. Spectinomycin/streptomycin can be successfully used for plant and bacterial...purpose="decorative" title="Check icon" %} Spectinomycin Excellent No Just bacteria Not SHuffle cells...

...base editor that converts adenosine to inosine in RNA. Since inosine is functionally equivalent to guanosine...to small indels in the target DNA that result in amino acid deletions, insertions, or frameshift mutations...enzymes. hfCas9s were developed by mutating specific amino acid residues to reduce off-target editing. Some... prime editing, and are often used to change an amino acid, turn a promoter on/off, or add small protein...with a silent mutation that does not change the amino acid sequence. HDR efficiency is generally low (...editors have been engineered to convert adenosine to inosine, which is treated like guanosine by the cell, creating...soon followed, using an enzyme evolved to excise inosine to access the same repair pathway and convert A...

...codons that encode 20 amino acids and three stop codons, meaning that the same amino acid can be encoded...actually prefer certain codons over others for certain amino acids. This is termed codon usage bias. In fact,...transfer RNAs (tRNAs). tRNAs bind to and deliver amino acids to the ribosome where they are incorporated...mRNA sequence. The 3’ end of  the tRNA binds the amino acid that corresponds to the anticodon sequence....always easy to identify the ideal codon for every amino acid across an entire polypeptide. Many researchers...wheel depicts the 64 codons that encodes the 20 amino acids and three stop codons. (B) Example of codon.... There are several codons that encode the same amino acid. Each organism, however, may preferentially...

...AraC system, which uses arabinose, requires many wash steps to remove arabinose making it difficult to ...🎶 “Bye bye L-arabinose drivePut your cultures in the shakerTurn the LEDs onAnd when you wantYou can ...recently published in @nchembio Title: Bye bye L-arabinose drive. Dedicated to Edo, Armin and @KhammashLab...lab,” says Di Ventura. It relies on the sugar arabinose to activate transcription by causing conformation...to regulate quickly. Arabinose also circulates within growth media and can’t be contained to a specific...AraC to make it responsive to light instead of arabinose. Nat Chem Biol. https://doi.org/10.1038/s41589...

...Wolf Frommer Reporter for arabinose levels in bacteria. The presence of arabinose enhances the FRET signal...14030 FRET Patrick Daugherty FRET: Vector for arabinose inducible expression of an N-terminally His-tagged...14031 FRET Patrick Daugherty FRET: Vector for arabinose inducible expression of an N-terminally His-tagged... LIC Series (8-Series) Various Plasmids pBAD (arabinose inducible, glucose repressible) His, Strep, mOCR...TEV cleavage sites to remove tags Scott Gradia Arabinose inducible vectors that come with various epitope...Various Plasmids pBAD, OR2-OR1-PR, pLtetO, pLlacO Arabinose, Anhydrotetracycline, lactose, IPTG Richard Murray...from the CFP and YFP fluorophores linked by an arabinose binding protein (AraF) in the plasmid. Other similar...

...autoinflammatory loop: keratinocytes both respond to and release IL-36 cytokines. Keratinocytes are the main source...cytokine family resulting in the activation of keratinocytes, which release inflammatory mediators that trigger...known how IL-36 cytokines actually get out of keratinocytes and induce their effector functions. It’s an...important to study what is happening within the keratinocytes. “We hope to use CRISPR libraries from Addgene...or proteins are involved in IL-36 release in keratinocytes,” says Mellett. This would help the lab identify...

...the ADAR2 deaminase convert adenosine to inosine on RNA. Inosine is translated as guanosine, so the overall...Cell . 154(2):442-51. PMID: 23849981 Ishino Y, Shinagawa H, Makino K, Amemura M, Nakata A. 1987. Nucleotide...genes in bacteriophages infecting Pseudomonas aeruginosa . Anti-CRISPR genes employ varied mechanisms ...MH, Morsut L, Liu Z, Brar GA, Torres SE, Stern-Ginossar N, Brandman O, Whitehead EH, Doudna JA, Lim WA...Abudayyeh OO, Gootenberg JS, Wu WY, Scott DA, Severinov K, van der Oost J, Zhang F. 2017. Multiplex gene...

...for aminoacyl tRNA synthetases By Kate Harten Recombinant antibodies targeting fourteen aminoacyl tRNA...escape in SARS-CoV-2 Recombinant antibodies for aminoacyl tRNA synthetases Nanocompartments for yeast bioproduction...pooled plasmid library encompassing all possible amino acid mutations of the receptor binding domain (RBD... work by generating antibodies for additional aminoacyl tRNA synthetases.     Figure 2: Specific...encapsulated a rate-limiting enzyme in this pathway (myo-inositol oxygenase), providing enzyme protection while ...

...Scaffold for inorganic phosphate biosensor Development of a Reagentless Biosensor for Inorganic Phosphate...1147-1155. Lin Tian Endocannabinoid GRAB_eCB2.0/eCBmut biosensor for endocannabinoids in vivo (Constitutive...CEPIA. Nat Commun. 2014 Jun 13;5:4153. Masamitsu Iino Calcium Red mitochondrial CEPIA calcium sensors ...mitochondria. Sci Rep. 2020 Feb 18;10(1):2835. Masamitsu Iino Calcium Red fluorescent calcium sensors (fRCaMP1...with an Expanded Dynamic Range by Optimization of Amino Acid Linkers between a Fluorescent Protein and PDE5alpha...sensor for spatiotemporally resolved imaging of endocannabinoid dynamics in vivo. Nat Biotechnol. 2021 Nov ... J Neurosci. 2006 Feb 15. 26(7):2000-9. Robert Malinow Endo/Exocytosis Synaptophysin-pHluorin Clathrin-mediated...