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Gut Check: using transfection to study the gut microbiome’s role in inflammation

Today’s SNiP reminds us that we are never truly alone. It’s been estimated that over one trillion microbial cells inhabit the human body. In this study, Wojciech et al. implicate a eukaryotic gut-dwelling microbe, Blastocystis subtype 7, in host T cell dysregulation, providing a mechanism for how this microbe can cause inflammation in the gut.

Generate Stable CHO Cell Lines

Using the CHOgro® High Yield Expression System, exceptional yields of recombinant protein can be obtained by transient transfection of suspension CHO cells. Though transgene expression ceases for the vast majority of cells within one to two weeks post-transfection, a small fraction of transfected CHO cells continues to stably express the recombinant protein through random chromosomal integration of the transfected gene. In this Tip from the Bench, we share a protocol for generating stable cell lines through selection and expansion of CHO cells which produce high levels of recombinant protein.

VirusGEN® and RoosterGEM™, a match made for GMP mRNA transfection

Here we highlight some of the exciting results from Mirus’ ongoing collaboration with RoosterBio. Earlier this year, RoosterBio presented a poster at ASGCT and ISCT which showcased using TransIT-VirusGEN® for high-efficiency transfection of mRNA into human mesenchymal stromal cells (hMSCs) cultured in RoosterGEM™ medium. Both VirusGEN® and RoosterGEM™ have GMP formulations available, which allows for rapid implementation into clinical manufacturing processes that require highly qualified reagents.

N to P Ratio

Cartoon of positive N divided by negative P.

In this TransMission Transfection 101, we bring you a brief primer on the ‘N to P ratio,’ which is a concept you may have encountered in the context of transfection. Here, we explain:

– What is the N to P ratio?
– How to calculate the N to P ratio
– Significance of the N to P ratio

Enrich CRISPR’d Cells with Label IT®

Icon showing cells sorted with Label IT and Cas9 RNP.

Today’s TransMission is a SNiP of one of our favorite papers featuring Label IT®. Nasri and Mir et al. describe an elegant use for Label IT® in enriching CRISPR/Cas-edited cells. For use in the clinic, it is highly desirable to limit the number of nonedited cells that may compete with the therapeutic, modified cells. Read on to learn how Label IT® was used to enhance the CRISPR genome-editing workflow!

What size are cell and gene therapies?

The relative size of AAV, lentivirus, transfection complex, and cell.

Have you ever given much thought to the relative size of the biological materials you work with every day in the lab? It can be mind-boggling to think about how something as tiny as a viral particle can have a life-saving impact for a patient with a debilitating genetic disease when delivered en masse.

But exactly how small of a package is all that life-changing power packed within? Test your knowledge in today’s blog post!

What’s in a (product) name?

What’s in a (product) name? That which we call a TransIT®
By any other name would deliver as efficient.

In today’s TransMission, we answer the FAQ: “Can I use this TransIT® to deliver that kind of nucleic acid?”

CRISPR Prime Editors Unleashed

“United we stand, divided we… small?”

CRISPR prime editing, introduced in 2019, harnesses a Frankensteinian enzyme–a Cas nickase fused to a reverse transcriptase–to perform gene edits with putatively higher fidelity than traditional CRISPR/Cas genome editing systems. In this SNiP, we highlight recent work from Grünewald et al. that shows the prime editor fusion can be split without negative consequence to editing, suggesting that the nickase and reverse transcriptase modules operate in trans. This finding is a boon for delivery and use of prime editing machinery with space-constrained vectors, such as AAV and lentiviral vectors.

Read on to learn more about prime editing and the authors’ discovery.

Transfecting Insect Cells, Pt. 4

Howdy everyone! What we have all BEEn waiting for: the final installment of our ‘Transfecting Insect Cells’ series with Dr. Bees.

So far, we have discussed common insect cell lines and TransIT®-Insect (Part 1), producing baculovirus with flashBAC™ (Part 2) and key features of pOET Transfer Plasmids (Part 3). Today in Part 4, we will bug out on how baculovirus can be used to produce virus-like particles (VLPs) and other kinds of viral vectors, such as AAV!