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  1. Viral Vectors 101: Producing Your rAAV

    Type
    Blog Post
    Published
    July 16, 2024, 1:15 p.m.
    ...ultracentrifuge. During the spin, the virions will form a band due to their high density, which can be collected...   Pro tip! An optional media exchange can be performed 12–18 hours post-transfection to help remove toxic...
  2. Hot Plasmids - November 2023

    Type
    Blog Post
    Published
    Nov. 14, 2023, 3:35 a.m.
    ...best PE6 variant for your application. Image reused form Doman and Pandey et al., 2023 under CC-BY 4.0 License.... These variants use different RT enzymes and performed large edits with similar or better efficiency ...potential to reduce negative side effects. This platform offers a promising approach for experimental therapies...we began distribution and has been verified to perform well in standard IHC and western blot protocols...protocols. You can find more specific information on the Wu Lab’s use of Anti-PSD-95 [K28/43R] under the Application...
  3. Plasmids 101: Choosing an Antibiotic Resistance Gene

    Type
    Blog Post
    Published
    Oct. 10, 2023, 1 p.m.
    ... plates. This prevents satellite colonies from forming while effectively selecting plasmid-containing ...cost-effective, timesaving for transformations. Cons: Prone to satellite colony formation on agar plates, less ...in a plasmid. During the propagation and/or transformation process, some bacteria will either never receive... is nifty because the recovery period after transformation (the dreaded 60 minutes shaking at 37 degrees..., interchangeable with Amp, timesaving for transformations. Con: Expensive.   Kanamycin Kanamycin, or...selection gene. However, it does require a slower transformation recovery period than amp or carb (60 minutes...confers mammalian resistance to G418. Con: Slow transformation   Spectinomycin/streptomycin Spectinomycin...
  4. Degrading DNA with Cascade-Cas3

    Type
    Blog Post
    Published
    Feb. 11, 2025, 2:15 p.m.
    ...mechanism (see next section for details). The newly formed single-stranded DNA (ssDNA) is then pulled through...through Cas3. After these initial steps, Cas3 can perform its helicase and nuclease functions simultaneously... D. W. (2023). Structural snapshots of R-loop formation by a type I-C CRISPR Cascade. Molecular Cell, ...
  5. Getting to Know Expansion Microscopy

    Type
    Blog Post
    Published
    June 20, 2023, 1:15 p.m.
    ...structure. For example, cytoskeletal structures that form continuous lines in pre-expanded samples can sometimes...
  6. Course-Based Undergraduate Research Experiences (CUREs)

    Type
    Blog Post
    Published
    Feb. 9, 2023, 2:15 p.m.
    ...partnered with colleagues at other institutions to form a support network called the MDH CUREs Community...). But is it possible to bring the kinds of transformative educational experiences afforded to those fortunate.... Therefore, he decided to adopt a more focused format, where all students work on malate dehydrogenase...hockey coach all shaped his teaching style, with the former offering excellent training in improvisation and...NADH production.   CUREs are beginning to transform undergraduate education in the sciences by making...
  7. CRISPR 101: Making a Knock-In Cell Line

    Type
    Blog Post
    Published
    Dec. 22, 2022, 2:15 p.m.
    ...location. HDR depends on resection at the break site to form ssDNA and uses the ends of the ssDNA to find a repair...increase HDR frequency by committing a break to performing HR before other factors were loaded onto the ...
  8. Tips for a 1st time AAV user (by a Rookie AAV user)

    Type
    Blog Post
    Published
    Oct. 23, 2018, 11:49 a.m.
    ...are required for proper packaging of AAV, but also form secondary structures that are prone to deletion ...and XmaI restriction sites). Another way is to transform AAV transfer plasmids into bacterial strains, ...found in the culture media, you should consider performing PEG precipitation. Instead of sonicating the ...
  9. Plasmids 101: Codon usage bias

    Type
    Blog Post
    Published
    Sept. 27, 2018, 1:09 p.m.
    ...choice, you should perform a functional assay to make sure your protein isn’t forming inclusion bodies ...
  10. CRISPR 101: Targeting RNA with Cas13a (C2c2)

    Type
    Blog Post
    Published
    Sept. 21, 2017, 2:07 p.m.
    ... in spacer acquisition following infection, as a form of “bait” to identify new CRISPR-associated proteins...target binding and cleavage through a series of conformational changes in the Cas13a molecule. Like Cas9, ...
  11. CRISPR Between the Genes: How to Experiment with Enhancers and Epigenomics

    Type
    Blog Post
    Published
    Aug. 9, 2016, 2:30 p.m.
    ...compaction—on the order of 700 terabytes per gram—and form the X-shaped chromosomes we all know and love. To...enhancers and genome math come in. They help us to transform raw genomic coordinates into meaningful experiments...tri_methylation In the next section we’ll apply the above formula to a real-world experiment. We’ll start with ChIP-seq...
  12. Minigenomes - a Safe Way to Study Dangerous Viruses Like the Ebola Virus

    Type
    Blog Post
    Published
    March 17, 2016, 2:30 p.m.
    ... associates with the viral polymerase, which is formed by a complex of two proteins: the large (L) catalytic...measured to quantify (indirectly) transcription performed by the viral polymerase. Alternatively, the mRNA...independently. In addition, experiments can be performed in the absence of some viral proteins (compared... minigenome plasmid per well (in a 6 well plate format). However, since plasmid quality and transfection...
  13. Avoiding the Dark Side of Fluorescent Protein Fusions with mOX FPs

    Type
    Blog Post
    Published
    Oct. 27, 2015, 3 p.m.
    ...and others have reported that FP cysteine residues form inappropriate disulfide bonds in the secretory pathway...modifications including glycosylation, disulfide bond formation, and proteolytic cleavage. FPs localized to organelles...fusion protein localization. For example, the formation of inappropriate inter-chain disulfide bonds between...because of the correct folding and fluorophore formation by most, if not all, of the FP fusions. Our moxFPs...
  14. Tips from the Repository Trenches: Using Barcodes to Track Samples

    Type
    Blog Post
    Published
    Oct. 6, 2015, 2:30 p.m.
    ...Barcoded tubes come in many forms. The 2D barcode can store a lot of information in a small space and is ... treated to easily take in plasmids (transformation). Transformed bacteria are then grown up on antibiotics...received is immediately transformed and stored. Some samples need to be transformed with a micropulser using... (98.2%) transform the first time and can be stored right away. Every week, the lab performs an average...average of 140 transformations. The lab typically transforms newly deposited samples from Monday through Wednesday...delivered by a mail courier and then the journey of transformation and storage begins. Some samples are submitted...barcoded tubes that are scanned into our Laboratory Information Management System (LIMS), a database we use to...
  15. Plasmids 101: Protein tags

    Type
    Blog Post
    Published
    Dec. 11, 2014, 4:26 p.m.
    ...can also protect against proteolysis. That’s one form of instability. Prokaryotes can also have a hard...Thioredoxin A (TrxA) that improve disulfide bond formation in order to help keep your protein soluble.  ... better assess their function. More detailed information and some protocols can be found in the references...
  16. Negotiating Work and Life: How to Find the Joy

    Type
    Blog Post
    Published
    March 20, 2014, 3:04 p.m.
    ... local family to help, create “fake” family by forming a social circle of people in the same situation... and/or your spouse won’t be at every school performance, chaperone every field trip or cheer at every...
  17. Plasmids 101: Plasmid Incompatibility

    Type
    Blog Post
    Published
    March 31, 2020, 1:15 p.m.
    ...separate plasmids bind to the same RepA molecules, forming a “handcuff” structure that physically blocks replication...provide clarity, a wet lab transformation experiment will ultimately inform whether your plasmids are ...about plasmids (Shintani et al., 2015). Now, bioinformatics tools help researchers determine how compatible...
  18. AAV Titers: Where Do They Come From And What Do They Mean?

    Type
    Blog Post
    Published
    Nov. 15, 2017, 3 p.m.
    ... of a viral genome). Empty viral capsids can be formed during viral production depending on the  experimental... the solution. These characteristics give us information on three important factors. What can AAV titers...requires an electron microscope, it is not routinely performed on all new viral vector preparations. Addgene ...
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