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Off-target Morpholino interaction reported based on mutant and 6ng dose

A prp1 Morpholino injected into zebrafish with mutated prp1 caused significant phenotype at a dose of 6 ng/embryo but had little effect at lower doses (see Supplemental Figure S13). This is a case where the Morpholino-in-mutant experiment not only detected an off-target interaction of the Morpholino but defined a dose where the effect suddenly became strong. Doses of prp1 Morpholino below that threshold produced phenotype at roughly the rate of the standard control oligo injections.

Tuesday, August 7, 2018 - 10:16
"Consensus guidelines for the use and interpretation of angiogenesis assays" with MO discussion

"Consensus guidelines for the use and interpretation of angiogenesis assays" contains a very good discussion of Morpholinos and specificity. Nowak-Sliwinska et al. extensively cite the Stainier et al. "Guidelines for Morpholino Use in Zebrafish". There is a theme in the Morpholino discussion in Nowak-Sliwinska et al.'s paper with which I disagree. They state "However, the best and generally accepted validation for any MO phenotype is confirmation of the same phenotype in a zebrafish genetic mutant." [1] Is this best, or does it exclude a valuable function of Morpholinos?

Thursday, May 24, 2018 - 12:02
Review (2008): Current perspectives in zebrafish reverse genetics

This is an older review (2008), but has a nice discussion of Morpholinos and controls. Photo-Morpholino and Vivo-Morpholino use in zebrafish are not addressed. The newer strategy of using Morpholinos in CRISPR mutants for specificity control is more recent than this paper, as is the understanding of gene compensation in mutants versus relatively uncompensated phenotypes from knockdowns triggered by Morpholino microinjections.

Monday, May 14, 2018 - 10:33
An i1e2 Morpholino triggering intron 1 retention: unusual splice outcome

Here is an example of an unexpected splice-modifying outcome. An oligo was targeted to i1e2, which would normally be expected to skip exon 2. Instead, the oligo caused retention of intron 1. This is the usual outcome of an e1i1 oligo, but not of an i1e2 oligo. Figure 5B shows the gel and the RNA map:

Friday, May 11, 2018 - 09:46
Splicing mutations in human genetic disorders: Review

This is an open-access review covering the mechanism of eukaryotic RNA splicing and diseases caused by mutations in regions affecting splicing.

Abramowicz A, Gos M. Splicing mutations in human genetic disorders: examples, detection, and confirmation. J Appl Genet. 2018. doi: 10.1007/s13353-018-0444-7

Tuesday, April 24, 2018 - 08:55
Antisense Phosphorodiamidate Morpholino Oligomers as Novel Antiviral Compounds (review)

Review article
Nan Y, Shang Y-J. Antisense Phosphorodiamidate Morpholino Oligomers as Novel Antiviral Compounds. Front Microbiol. 2018;9:750. doi:10.3389/fmicb.2018.00750

Monday, April 23, 2018 - 07:33
BLAST homology: screening predicted specificity

It is always prudent to screen proposed oligo sequence for potential off-target RNA interactions using BLAST. Partial homologies between a Morpholino's inverse complement and an RNA which are reported by BLAST are often in regions of the RNA where binding a Morpholino is unlikely to alter gene expression. If there is significant homology in a region where binding a Morpholino is likely to alter a transcript's expression, have us design another oligo.

Thursday, April 5, 2018 - 09:31
Fish mutant, where is thy phenotype?

"Due to genetic compensation, phenotypically wild-type mutants can become refractive to morpholino-induced phenotypes, providing a critical test both for genetic compensation and for the specificity of morpholino phenotypes."

Balciunas D. Fish mutant, where is thy phenotype? PLoS Genet. 2018;14(2):e1007197.

Thursday, February 22, 2018 - 11:40
Cardiac Ventricular Injection in Zebrafish Larva

This is an interesting technique for later-stage zebrafish embryo treatment, involving pulsed injection of a mixture of Morpholino and Endo-Porter into the heart. I think this might push the limit of solubility for many Morpholino sequences. They are reporting good systemic delivery -- see the video for fluorescent images of the oligo distribution.

Wednesday, February 14, 2018 - 15:27
Patent on central nervous system delivery of Morpholinos by low osmolar contrast agents

This one looks interesting. Morpholino and 2'-MOE activities are compared in SMA models.

United States Patent Application 20180030443
February 1, 2018
Inventors: Burghes; Arthur; (Columbus, OH) ; Porensky; Paul; (Worthington, OH) ; Kaspar; Brian; (New Albany, OH)


Wednesday, February 7, 2018 - 11:43