...makes polyclonal antibodies unique and what experiments you should choose them for. How polyclonal antibodies...cross-reactivity. When to use polyclonal antibodies One advantage of polyclonal antibodies is that they are...barrier to entry makes polyclonal antibodies a common reagent in many labs. Polyclonal antibodies also have...When to stay away from polyclonal antibodies The most prevalent issue with polyclonal antibodies is their...from the same species your polyclonal antibody was generated in or a polyclonal isotype control, to account...options to choose from. In this article we’ll cover polyclonal antibodies, one of the many different types of...antibodies are produced Polyclonal antibodies are a heterogeneous mixture of many antibodies that recognize...

...pathophysiological role of glycans. Follow her on Facebook: @glycoimmunologygroup.     Michael McMurray is...Addgene to advance their research on diseases of glycosylation, arginine mutagenesis, and neural development...development. Paula Videira: Congenital disorders of glycosylation Paula Videira is an Assistant Professor at the...University in Lisbon, Portugal. She leads the Glycoimmunology group based at the Research Unit on Applied...physiological and pathological roles of human cell glycans. Accordingly, she joined efforts with the Portuguese...Portuguese Association for Congenital Disorders of Glycosylation (APCDG) and an international platform for CDG...CDG&Allies-PPAIN).  Congenital disorders of glycosylation (CDG) are a group of rare inherited metabolic...

... CRISPR flyCRISPR Lab CRISPR Plasmids flyCRISPR Plasmids Available from Addgene... Wildonger Lab - O'Connor-Giles Lab email: flycrispr@gmail.com We have found that a variety of Cas9...Resources: Protocol for generating gRNAs 106.5 KB flyCRISPR website contains additional information CRISPR...

...Creating Bacterial Glycerol Stocks for Long-term Storage of Plasmids Creating Bacterial Glycerol Stocks for Long-term...time, you will need to establish glycerol stocks. The addition of glycerol stabilizes the frozen bacteria...of 50% glycerol in a 2 mL screw top tube or cryovial and gently mix. Notes: Make the 50% glycerol solution...concentration of long-term glycerol storage is unknown. Most labs store bacteria in 15-25% glycerol. You can prepare...your glycerol stock before you begin your plasmid mini-prep. Try not to freeze/thaw your glycerol stock...Protocol for creating a glycerol stock, including additional tips for long-term storage. Protocols...Bacterial Culture Background Information Bacterial glycerol stocks are important for long-term storage of ...

... N-methylpurine DNA glycosylase (MPG), also known as alkyladenine DNA glycosylase (AAG). When adenine ... MPG / AAG N-methylpurine DNA glycosylase / alkyladenine DNA glycosylase  Removes hypoxanthine from DNA...using either fused or endogenous Uracil DNA N-glycosylase (UNG) enzymes. Various other proteins included... (Y = T or C) base editor, which they called glycosylase-based guanine base editor (gGBE) (Tong, Liu, ...Editor Converts C → G by fusing a uracil DNA glycosylase to a CBE; the first type of base transition editor...apyrimidinic (AP) site. UNG / UDG Uracil DNA N-Glycosylase Removes uracil from DNA, creating an abasic site...base), or C, respectively. gGBE, gTBE, gCBE Glycosylase-based G, T, or C base editors Excises G, T, or...

... use a different envelope glycoprotein instead of the wild-type glycoprotein. The pseudotyped protein ... on lentiviral particles. Rabies glycoprotein The rabies glycoprotein targets neuronal cells and can travel...neurons that express TVA and the rabies glycoprotein. The glycoprotein will be incorporated into the replicating...sense to pseudotype another virus with rabies glycoprotein. Lentiviral vectors are also great at infecting...peripheral gene delivery. Combining the rabies glycoprotein into the lentiviral envelope allows the lentiviral...alone.  This pseudotyped lentivirus with rabies glycoprotein now serves as a retrograde tracer so that scientists...Another way to use rabies virus is to delete the glycoprotein required for entry into the cell (rabies dG)...

...that they called a Glycosylase Base Editor, or GBE, for its reliance on the glycosylase UNG to crease abasic...Converts C → G; also called a glycosylase base editor (GBE). GBE Glycosylase Base Editor A base editor ...Uracil DNA N-Glycosylase Removes uracil from DNA, creating an abasic site. UGI Uracil Glycosylase Inhibitor...at its original job. They removed the uracil glycosylase inhibitor (UGI) components, which inhibit BER...instead introduced its opposite, a uracil DNA N-glycosylase (UNG) from E. coli. The resulting construct, ...editor that relies on DNA glycosylation to create an abasic site. In this post, it refers to the same approach..., Rosser, S. J., Bi, C., & Zhang, X. (2021). Glycosylase base editors enable C-to-A and C-to-G base changes...

...bacteria, or monoderms, have a single thick peptidoglycan membrane. Common gram positive bacteria include...negative bacteria, or diderms, have a thinner peptidoglycan membrane and an outer lipid membrane. Common...salmonella and E. coli. Because of their thicker peptidoglycan layer, gram positive bacteria trap the crystal...violet (CV) is water soluble and can enter the peptidoglycan layer of both bacteria types. In water, the ...with the CV+, creating a complex within the peptidoglycan layer, staining the cell purple. When the alcohol...alcohol is added, it dehydrates the peptidoglycan layer, trapping the large dye complex created by the...the outer lipid membrane, and the thin inner peptidoglycan membrane is unable to retain the stain, leading...

...protein products, specifically glycosylation. The native glycosylation machinery of K. phaffii lacks certain...source Glucose Glucose Glucose, methanol, glycerol Heterothallic mating Efficient Efficient...source of carbon and energy (as well as glucose or glycerol). K. phaffii is only distantly related to S. pombe... certain terminal mannose glycopeptides (which most other yeast possess) that are problematic for human...human administration. Thus, the production of glycoproteins for human therapies, such as antibodies, is ...in K. phaffii since it produces ‘human-like’ glycosylated proteins. Peroxisome studies Peroxisomes are...

...sequencing is done from glycerol stocks (overnight cultures with 15% glycerol), as opposed to purified...for glycerol stock production and storage, we grow a little extra and transfer 100 µl of glycerol stock...uniformity of the culture. Sequencing directly from the glycerol stock saves us the step of purifying the DNA and...and then shipped along with their corresponding glycerol plate on dry ice to the sequencing provider. Each...are equipped to perform Sanger sequencing from glycerol stocks, so make sure to select the appropriate...

...lot number while you’re at it! Polyclonal or monoclonal Both polyclonal and monoclonal antibodies can ...conditions Pros Cons Tris-glycine 6–400 kDa Tris-glycine 100 V, 1–2 hours Easy and cheap...can work well for a western blot. Polyclonal antibodies, which bind to multiple sites on an antigen, are...protein — can be used to reproduce the benefits of polyclonal antibodies. Because these mixtures are defined...are not as commonly available as monoclonal or polyclonal antibodies, but it’s worth a look to see if they...Precast gel Increased cost Save money Polyclonal antibodies Higher non-specific binding; lot-to-lot...available; cost varies widely Increase signal Polyclonal antibodies Higher non-specific binding; lot-to-lot...

...gRNAs. gRNAs are flanked by glycine tRNAs to create polycistronic glycine tRNA-gRNA (PTG) constructs. ...transcript You can also multiplex gRNAs via a polycistronic transcript. Rather than being transcribed from...option described below, plasmids for making polycistronic gRNAs are also available from the Yang lab for...Plasmids: These plasmids are similar to the Csy4 polycistronic system described above, except that they use...deaminase fusion with Cas9 with a uracil DNA glycosylase inhibitor with a degradation tag (LVA tag) to...

...gRNA cleavage.   Xie et. al constructed polycistronic glycine tRNA-gRNA genes (PTGs) in the forms tRNA-gRNA...separated by 28 base Csy4 recognition sites from a polycistronic transcript. Although this method shortens construct...requires coexpression of Csy4 with Cas9 and the polycistronic transcript. As nucleases may have off-target...Bacteria and viruses have illustrated the power of polycistronic gene organization, and its translation to CRISPR...

...question you may have is whether a monoclonal or polyclonal antibody is better for your experiment. Monoclonal...antibody clone and therefore a single isotype while polyclonal antibodies contain a mix of different isotypes...tend towards more reproducible results. Since polyclonal antibodies contain a variety of clones and thus...secondary antibodies instead (B).   Since a polyclonal primary antibody contains a mix of different ...reactive secondary antibodies when you’re using polyclonal primary antibodies.  I hope this post has explained...

...that encode N-glycosylation consensus sequences are modified with the addition of N-glycans [9]. Both of... post-translational modifications including glycosylation, disulfide bond formation, and proteolytic cleavage...re-engineered by mutating cysteine residues and N-glycosylation consensus sequences in popular FPs. This generated...Costantini, Lindsey M., et al. "Cysteineless non-glycosylated monomeric blue fluorescent protein, secBFP2,...

...to express the transcription factors from a polycistronic cassette. However, it was unknown whether variations...variations in gene expression from these polycistronic cassettes could lead to comparable experimental...Woltjen Lab compared different variants of the polycistronic cassettes and discovered reprogramming discrepancies...variation in the Klf4 isoform (KLF4s or KLF4L). Polycistronic cassettes that contained KLF4s had overall diminished...reprogramming and global gene expression patterns. Polycistronic cassettes that contained the KLF4L isoform had...the observed reprogramming discrepancies. The polycistronic piggyBac transposon vectors tested in this work...

...understood. Much of the action depends on a surface glycoprotein – the only viral protein present on the virion...in PLoS Pathogens shows that the Ebola virus glycoprotein remains essential in allowing those infected...Medicine have also recently described how the glycoprotein transforms itself to escape vesicles within ...reported in the Journal of Virology.  The EBOV glycoprotein’s role in infection also makes it a critical ...

...target protein, so they are referred to as polyclonal. Polyclonal antibodies have the advantage of more robustly...find more details about polyclonal antibodies on our Antibodies 101: Polyclonal Antibodies blog post. Monoclonal...advantages and disadvantages compared to each other. Polyclonal antibodies These antibodies are derived from ...

...Action*   Working Concentration** Kanamycin aminoglycoside Binds 30S ribosomal subunit; causes mis-translation...mis-translation Bactericidal 50-100 ug/mL Spectinomycin aminoglycoside Binds 30S ribosomal subunit; interrupts protein... Bactericidal 7.5-50 ug/mL Streptomycin aminoglycoside Inhibits initiation of protein synthesis ...synthesis Bactericidal 100 ug/mL Bleomycin glycopeptide Induces DNA breaks Bactericidal 5-100 ug/mL...

... be shared across other proteins. Conversely, polyclonal antibodies - which are a heterogenous mixture...antibodies, their cost is higher than that of polyclonal antibodies. For example, it takes longer to develop...than it does to immunize an animal and recover polyclonal antibodies (Lipman et al., 2005). This is reflected...Trudel LJ, Weis-Garcia F (2005) Monoclonal Versus Polyclonal Antibodies: Distinguishing Characteristics, Applications...