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Using Morpholinos to Control Gene Expression (in CPNAC)

Wiley has published an update to my Morpholino how-to and background review.

Moulton JD. Using Morpholinos to Control Gene Expression. Curr Protoc Nucleic Acid Chem. 2017 Mar 2;68:4.30.1-4.30.29. doi: 10.1002/cpnc.21.

Saturday, March 4, 2017 - 03:26
Fructose and uptake of Morpholinos

This is about fructose and Morpholinos.

Here is the original paper being discussed.

Friday, March 3, 2017 - 08:58
nasal administration of delivery-enabled Morpholino in mice

This paper is an example of nasal administration of a delivery-enabled Morpholino in mice.

Soonthornvacharin S, Rodriguez-Frandsen A, Zhou Y, Galvez F, Huffmaster NJ, Tripathi S, Balasubramaniam VRMT, Inoue A, de Castro E, Moulton H, Stein DA, Sánchez-Aparicio MT, De Jesus PD, Nguyen Q, König R, Krogan NJ, García-Sastre A, Yoh SM, Chanda SK. Systems-based analysis of RIG-I-dependent signalling identifies KHSRP as an inhibitor of RIG-I receptor activation. Nat Microbio. 2017;2:17022. doi:10.1038/nmicrobiol.2017.22

Friday, March 3, 2017 - 06:55
Review: Eteplirsen in the treatment of Duchenne muscular dystrophy

Lim KRQ, Maruyama R, Yokota T. Eteplirsen in the treatment of Duchenne muscular dystrophy. Drug Design, Dev & Ther. 2017;11:533-45. doi:DDDT.S97635

Open access

Wednesday, March 1, 2017 - 08:06
Abdominal injection of Vivo-Morpholino in fish for oocyte knockdown

Injection of Vivo-Morpholinos into the abdominal cavity of medaka was used to knock down mPRα in the oocytes.

Roy SR, Wang J, Rana MR, Nakashima M, Tokumoto T. Characterization of membrane progestin receptor α (mPRα) of the medaka and role in the induction of oocyte maturation. Biomed Res. 2017;38(1):79-87. doi: 10.2220/biomedres.38.79.

Wednesday, March 1, 2017 - 04:04
TEDx Perth talk on Morpholinos and Duchenne muscular dystrophy (Wilton, late 2016)

Prof. Steve Wilton: TEDx Perth talk on Morpholinos and Duchenne muscular dystrophy.

If you have had time with Steve, life is better. He's a good sort.

Thursday, February 16, 2017 - 01:12
Which splice junction to target, predicting cryptic splice sites, and doing the two-non-overlapping oligo specificity experiment

I've had the pleasure of corresponding with Yuhong Liang (梁雨虹)of 4A Biotech in China; she is the product manager for Morpholino oligos at 4A Biotech. We have been working with 4A for a little over a year now and Yuhong has been learning very quickly, but is still new enough to the Morpholino field that she is asking questions of broad interest. She had some questions regarding which splice junction to target, predicting cryptic splice sites, and the process of doing the two-non-overlapping oligo specificity experiment.

Wednesday, February 8, 2017 - 01:07
Getting Morpholinos into cultured cells

There are three usual options to consider for cell culture experiments with Morpholinos. There are other methods, but most labs choose one of these.

(1) If your cells can tolerate electroporation, that is an inexpensive option for delivery of standard Morpholino oligos. However, electroporation causes high cell mortality for many cell types.

Friday, December 16, 2016 - 04:44
Validating Morpholino phenotypes with CRISPRs

I've been enjoying a conversation with Martin Blum about Morpholinos, CRISPR mutants, funding and publication. He wrote:

Good morning Jon,

My study section at the German Research Council DFG now accepts MO-proposals, no problem. Still, many people particularly working in zebrafish are too careless with controls and rescues. That might backfire at one point in the future.

My take is that we will use CRISPR to validate MOs and then use MOs most of the time.

Best wishes,

Thursday, December 8, 2016 - 04:37
Assessing activity of a splice-modifying Morpholino

This is about detecting the activity of a splice-modifying oligo by reverse-transcriptase PCR and gel electrophoresis.

The expected outcome of an exon targeting an exon 3 splice junction (either i2e3 or e3i3) is elimination of exon 3 from the mature mRNA (along with both of its flanking introns, i2 and i3). The typical way to assess activity of a splice-modifying oligo is to isolate RNA from treated samples and run reverse-transcriptase PCR (RT-PCR), then run the PCR product on a gel to assess the mass of the product against a DNA ladder.

Friday, December 2, 2016 - 03:49