... two global stock centers for Arabidopsis thaliana (Arabidopsis). Our mission is to collect, preserve,...in 60 countries each year. Introducing Arabidopsis Arabidopsis is an important model system for research...and you have yourself a plant publicity crisis. The Arabidopsis Biological Resource Center (ABRC) set out...Translating Research on Arabidopsis Into a Network of Educational Resources). Arabidopsis is used in research... the Education & Outreach Specialist for the Arabidopsis Biological Resource Center and the Center for...Ohio State University. Established in 1991, the Arabidopsis Biological Resource Center (ABRC) is one of two...distribute seeds, DNA and other resources for Arabidopsis and related species. Located at The Ohio State...

...spectinomycin resistant phenotype in bacteria. While there are many known spectinomycin resistance mutations...genomic site for mutagenesis, a feature which they used to generate bacteria resistant to both spectinomycin...directed mutagenesis to generate specific mutations in your plasmids Use REPLACER Mutagenesis to mutate...Mutagenesis is a tool that both evolution and molecular biologists use to tinker with DNA. Making changes...region. EvolvR, a CRISPR-Cas9 based targeted mutagenesis method developed by the Dueber Lab at Berkeley...potential applications. Challenges of existing mutagenesis methods Several continuous diversification methods...error prone polymerase I (PolI3M) is capable of mutagenesis in E. Coli, but this approach requires special...

..., a graduate student at Stanford University. Arabidopsis is a fantastic model organism for many reasons... are many motivations to generate transgenic Arabidopsis, from studying transcriptional and translational... plants, to complementing mutant phenotypes. Arabidopsis is amenable to the floral drip or dip transformation...transformation protocol, as well as protocols for growing Arabidopsis are also available (Weigel and Glazebrook, 2002...vector is the first step in generating transgenic Arabidopsis. There are a bunch of great options, but here... pool you will be selecting from later. The Arabidopsis transformation process The process itself is ... the chances of tDNA integration. Selecting Arabidopsis transformants Once your seeds from your T0 plants...

... the mutagenesis reaction. Alternative approaches to site directed mutagenesis Point-mutagenesis is fairly...constructs), in functional analysis of proteins (to perform alanine scanning mutagenesis or targeted substitution...should be resistant to Cas9 cleavage. If a plasmid contains the template, site-directed mutagenesis can be...resulting construct resistant to Cas9 induced cleavage. Summary of Site Directed Mutagenesis In brief, point-mutations...mutation of interest. Gel electrophoresis diagrams of the anticipated restriction analysis results (left) and...Laursen from Cornell University. Site directed mutagenesis is a highly versatile technique that can be used...substitution of key residues), and in SNP analysis (to introduce naturally occuring SNPs in a plasmid context...

...protein synthesis Bactericidal 25-100 ug/mL Ampicillin beta-lactam Inhibits cell wall synthesis Bactericidal...Resistance to antibiotics is a widely used tool in molecular biology, yet scientists rarely stop to think...to maintain the plasmid. Adding an antibiotic resistance gene to the plasmid solves both problems at once...pressure to keep your plasmid. Viva la (bacterial) resistance!  What are antibiotics? Antibiotics are generally...Binds 30S ribosomal subunit; interrupts protein synthesis Bactericidal 7.5-50 ug/mL Streptomycin aminoglycoside...Carbenicillin beta-lactam Inhibits cell wall synthesis Bactericidal 100 ug/mL Bleomycin glycopeptide...Binds 30S ribosomal subunit; inhibits protein synthesis (elongation step) Bacteriostatic 10 ug/mL ...

...Microbiology Pages What is antibiotic resistance? Antibiotic resistance is defined as microbial survival despite...generally organisms with multi-drug resistance are resistant to three or more different classes, or types...Learn How Antibiotic Resistance Genes Are Used in the Lab How does antibiotic resistance develop? There are...in which bacteria can become resistant to antibacterial drugs. Resistance can be intrinsic, meaning a ...means the bacterium is not naturally resistant but has become resistant somehow overtime. One fantastic example... example of intrinsic resistance would be Mycoplasma species’ resistance to beta-lactam antibiotics, like...Mycoplasma are intrinsically resistant to this class of antibiotic. Acquired resistance can be the result of ...

...beta-lactam that inhibits cell wall synthesis. The AmpR (amp resistance) gene produces beta-lactamase to ...processes (e.g., protein synthesis), while amp only affects the cell wall synthesis step during cell division...aminoglycoside which inhibits protein synthesis. The kan resistance gene, NPTII, facilitates phosphorylation...cells, are already resistant to this antibiotic due to stable integration of resistance genes in their genomes...damage. Choosing an antibiotic resistance Choosing the best antibiotic resistance ultimately comes down to ...Plasmids need antibiotic resistance genes so that you can specifically isolate bacteria containing your...interest. But does it matter which antibiotic resistance gene you select? In this blog, we’ll break down...

...this insertion size, primer synthesis is more expensive than gBlock synthesis, and multifragment assembly...REPLACR-mutagenesis could save busy scientists a lot of time. Let us know how REPLACR-mutagenesis works ...Comparison of REPLACR-mutagenesis with other common methods for site directed mutagenesis   References 1. ...al. (2016). “REPLACR-mutagenesis: a one-step method for site-directed mutagenesis by recombineering.” ...Site-directed mutagenesis (SDM) is one of the key tools researchers use to prove causation in molecular...vitro assembly methods. A new method, REPLACR-mutagenesis, harnesses the power of bacterial recombineering... Plasmids After PCR). Current site directed mutagenesis methods Current SDM methods rely on in vitro ...

...interest, C-terminal tags, plant terminators and resistance cassettes for selection of transgenic plants....Lohmann said they’ve only tested GreenGate in Arabidopsis, it should in principle work in any plant species...Versatile and Efficient Cloning System for Plant Transgenesis." PLoS ONE. 8(12): e83043 (20 December 2013)...

...Apoptosis or “programmed cell death" plays a pivotal role in an array of biological processes including...development, the immune system, and cell turnover. Apoptosis is a highly controlled process that is triggered...fragmentation and death (Elmore, 2007). To study apoptosis in vivo, scientists have used genetically encoded...fluorescent executioner caspase reporters. Several apoptosis reporters have been developed, such as the Forster...Zhang et al., 2019). Thus to effectively study apoptosis in vivo scientists need a protease reporter that...brightness. The Shu Lab from UCSF designed an improved apoptosis reporter, ZipGFP, which utilized the self-assembling... better study the spatio-temporal dynamics of apoptosis in living animals. To do this they modified a ...

...effects. One-pot saturation mutagenesis The one-pot saturation mutagenesis technique described in Wrenbeck... as the gene or use custom gene synthesis or site directed mutagenesis to remove the BbvCI site(s). ...saturation mutagenesis with the same protocol. What do you think about one pot saturation mutagenesis? How .... “Random Mutagenesis by Error-Prone PCR”. Methods in Molecular Biology In Vitro Mutagenesis Protocols...the Whitehead lab developed One pot saturation mutagenesis as a quick and easy technique that can be used...library generation prior to one pot saturation mutagenesis Several methods can be used to create mutational... shortcomings compared to one pot saturation mutagenesis. Error-prone PCR: This method amplifies the...

...take a look at our live examples: experiment or analysis. In the rest of this article, we'll explain the...batch guide design, batch primer design or batch analysis. We wanted to relieve scientists of the burden... by Luca Pinello and Kendell Clement for batch analysis because of its ease-of-use and comprehensive reports...publicly-available for high-throughput design and analysis of CRISPR experiments. It has certainly boosted...

...research on diseases of glycosylation, arginine mutagenesis, and neural development. Paula Videira: Congenital...disorders of glycosylation Paula Videira is an Assistant Professor at the Faculty of Sciences of NOVA University...functional activity of VUSs will enable more accurate diagnosis of NFIX-related syndromes (Sotos-type 2 and Marshall-Smith...

...plasmids. Here, we'll cover restriction digest analysis. Diagnostic restriction digest Diagnostic digests...different features within the plasmid. Restriction analysis can also be used successfully even if you don'...

... more long-term solution is the use of unique persistent identifiers (PIDs), a long-lasting way to identify...documents, files, or physical reagents. What are persistent identifiers? Perhaps the most commonly known ...original article online. It is also designed to be persistent and exist online indefinitely. PIDs historically...plasmids! In an effort to support standardized persistent identification of research reagents, several ...this post and sharing her technical expertise in persistent identifiers. Additional Resources on the Addgene...

...Mycobacterium tuberculosis resistance to pyrazinamide (PZA) (a drug commonly used to treat tuberculosis) and in...projects.   Understanding drug resistance mechanisms of Mycobacterium tuberculosis Dr. Patricia Sheen, Universidad... accurate detection of PZA-resistance is necessary to improve tuberculosis care and control. In her current...methods and improved therapies for PZA-resistant Mycobacteria tuberculosis. Investigation of multispecies biofilm...Sheen aims to identify new genes related to PZA-resistance by using different approaches, including CRISPR...experience in the field of bacterial and fungal pathogenesis. Her research focuses on a collection of pathogenic...

... Protocols Sequence Analysis Sequence Analysis of a Plasmid You ...performing sequence analysis of a plasmid Protocols...

...Nishimasu H, Ran FA, Hsu PD, Konermann S, Shehata SI, Dohmae N, Ishitani R, Zhang F, Nureki O. Cell. 2014...transcriptional activation of endogenous genes and it consists of three components: 1. A nucleolytically inactive... a synthetic transcription activation complex consisting of multiple distinct effector domains that work...genes that, when turned on, make melanoma cells resistant to treatment with a BRAF inhibitor. (The test ...-mediated screens exhibited a high degree of consistency and validation with >80% effectively enriched...

...only measures infectious viral particles. One of the si mplest methods of measuring functional titer uses...components such as LTRs, gag, WPRE, antibiotic resistance-genes, or the transgene itself. Many researchers...Another popular approach utilizes the antibiotic resistance genes carried on transfer plasmids. Target cells... one copy integrates, selecting by antibiotic-resistance or single cell sorting based on expression of...

...Nishimasu H, Ran FA, Hsu PD, Konermann S, Shehata SI, Dohmae N, Ishitani R, Zhang F, Nureki O (2014) Crystal...