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Splice modulation and exon skipping ASOs

AUMsplice sdASO

Self-delivering antisense oligonucleotides for splice modulation and exon skipping

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Inside the nucleus at the top right of a cell, a self-delivering antisense oligonucleotide, one uniform strand with no gap, is paired with one exon of a pre-messenger RNA, and that exon hangs in a loop below the spliceosome, a large translucent complex of several rounded bodies, which holds the two flanking exons together above it under a warm glow, so the looped exon is skipped; the oligonucleotide stays whole and the RNA it binds is not cut, with no enzyme on it. Chromatin fills the nucleus, another strand passes through a nuclear pore in the envelope, and in the cytoplasm below single strands drift, one leaving a vesicle, while the cell membrane takes one in from the medium through a pit, with no transfection reagent.

AUMsplice sdASOs are self-delivering antisense oligonucleotides (ASOs) for splice modulation. They bind pre-mRNA and redirect splicing, typically by causing a specific exon to be skipped (left out) during mRNA processing. By skipping a targeted exon, AUMsplice sdASOs allow researchers to alter protein coding sequences or restore reading frames in cases of mutations. They are also used to study the functional importance of exons. AUMskip sdASO is the exon-skipping product within AUMsplice sdASO, and it is what most researchers ask for.

Splicing is redirected in the nucleus, and many of the cells where that is studied are difficult to transfect with conventional methods. Transfection reagents, electroporation, or viral vectors can introduce toxicity, alter the cellular response, or require additional optimization. AUMsplice sdASOs enter the cell without a transfection reagent, electroporation, or a viral vector. They are built on the AUMsilence platform, whose chemical modifications enable self-delivery. Add them to the culture medium or inject them into an animal. Like AUMblock sdASO, AUMsplice sdASO belongs to the steric-blocking category – it modulates RNA splicing through binding and blocking splice sites, without eliciting RNase H cleavage of the RNA.

Sizes and purification

Available by quote: tell us your target and we design and price it.

Prices and lead time are shown on the order page. Custom sizes by quote.

At a glance

Class
Self-delivering antisense (sdASO)
Targets
Splice modulation and exon skipping
Delivery
Add to culture medium or inject in vivo.
Fluorescent label
None

In the lab

The protocol in four steps

The steps below are the AUMsplice sdASO protocol's, in its own words. The full protocol carries the amounts per plate format, the tips and the troubleshooting.

Read the full protocol

  1. Step 1

    Cell preparation

    Plate cells in their optimum growth medium at a density appropriate for the cell type. A confluency of 50-70% at the time of treatment is typical.

  2. Step 2

    AUMsplice sdASO stock preparation

    Prepare AUMsplice sdASO stock solution by reconstituting lyophilized ASOs at the desired concentration. If you already have a stock solution prepared, skip to Step 3.

  3. Step 3

    AUMsplice sdASO delivery to cells

    Add AUMsplice sdASO to the cells at the desired final concentration. The recommended working range is 5-20 μM, with a starting concentration of 10 μM. The optimal concentration varies with the target gene, the RNA class (messenger RNA, microRNA or long non-coding RNA) and the cell type, and should be determined by titration for each system.

  4. Step 4

    Incubation and analysis

    Incubate cells with AUMsplice sdASO and analyze splice modulation at appropriate time points.

FigureThe workflow in cell culture, in four steps

In vitro, the four steps

  1. Step 1

    Prepare cells

    Seed the cells in a 24-well plate; the oligonucleotide waits in solution.

  2. Step 2

    Add to the medium

    Add the self-delivering oligonucleotide straight to the medium: no transfection reagent, no carrier.

  3. Step 3

    Incubate

    Incubate at 37 degrees C in 5% carbon dioxide, humidified, for the exposure time.

  4. Step 4

    Quantify knockdown

    Measure the knockdown: RT-qPCR, immunoblot, flow cytometry.

    Schematic of the three readouts, not data.

For research use only. Not for use in diagnostic or therapeutic procedures.
Sources
  1. The pictures are the object pictures of AUM BioTech's workflow slide, rendered views of the objects on a white ground; none is a photograph of an experiment and none records a measurement.
  2. Tile 04 is a schematic of the three readouts, not data: the two conditions are drawn with a modest difference to show what each readout is, and no magnitude is asserted.

Why

Features of AUMsplice sdASO

  • AUMskip sdASO: the exon-skipping application of AUMsplice sdASO

    AUMskip sdASOs are designed to target splice sites or splicing regulatory elements, causing the spliceosome to bypass specific exons during pre-mRNA processing. This results in the production of mRNAs lacking those exons, which can translate into truncated but potentially functional proteins.

  • Disease-relevant applications

    Exon skipping is a proven strategy for addressing certain genetic diseases. For example, in Duchenne Muscular Dystrophy, skipping mutated exons can restore the reading frame of the dystrophin gene, allowing production of a shortened but partially functional protein. AUMsplice sdASO enables research in this area.

  • Self-delivery to the nucleus

    AUMsplice sdASOs are designed to be taken up by endocytosis without transfection reagents and to reach the nucleus, where pre-mRNA splicing occurs. They are used to modulate splicing in primary cells, tissues, and in vivo models that are difficult to transfect with conventional methods.

  • Sequence design against the target exon

    Every AUMsplice sdASO is designed by AUM against the target the customer names, as part of the price. Several sequences are designed against different sites in and around the target exon, since which one works best is not predictable in advance. Each is checked for specificity against the transcriptome before synthesis.

Mechanism

Mechanism of action

Splice site blocking: After the AUMsplice sdASO is delivered into the nucleus of a cell, it binds to its pre-mRNA target at or near the exon one intends to skip. Common binding sites include the 3′ splice acceptor site, 5′ splice donor site, or splicing enhancer sequences.

Spliceosome redirection: When the spliceosome assembles on the pre-mRNA, the bound AUMsplice sdASO sterically hinders the binding of splicing factors that normally recognize the exon's boundaries. Essentially, the ASO "hides" the exon from the splicing machinery.

Altered mRNA production: As a consequence, the spliceosome skips over that exon, splicing together the flanking exons (e.g., exon N joined to exon N+2, leaving out exon N+1). The outcome is the production of an mRNA that is missing the targeted exon, resulting in a modified protein product.

FigureHow AUMsplice sdASO acts
Three stages inside the nucleus, left to right: one uniform oligonucleotide strand with no gap pairs on the middle exon of a capped pre-messenger RNA that runs to a beaded tail; the spliceosome, a large translucent complex of rounded bodies, holds the two flanking exons together at one junction under a warm glow while the paired exon and its introns hang below it as a single teardrop loop; the mature message, with one cap and one tail, leaves cap first through the middle of three pore complexes in the nuclear envelope at the right, and the released loop, closed on itself with the strand still paired, drifts below. No enzyme appears and nothing is cut; chromatin fills the background.
  1. Stage 1

    AUMsplice sdASOs are designed to be taken up by endocytosis without transfection reagents and to reach the nucleus, where pre-mRNA splicing occurs.

  2. Stage 2

    After the AUMsplice sdASO is delivered into the nucleus of a cell, it binds to its pre-mRNA target at or near the exon one intends to skip.

  3. Stage 3

    The bound AUMsplice sdASO sterically hinders the binding of splicing factors that normally recognize the exon's boundaries. As a consequence, the spliceosome skips over that exon, splicing together the flanking exons.

Applications

Applications and use cases

  • Genetic disease research

    AUMsplice sdASO is used in research on diseases like Duchenne Muscular Dystrophy (DMD), where skipping a mutated exon can restore the dystrophin reading frame. It is also used in research on Spinal Muscular Atrophy (SMA), where the splice-modulation strategy is exon inclusion rather than skipping. AUM designs the sequences against a patient-specific mutation or against a general disease model.

  • Alternative splicing research

    If you want to study what an alternatively spliced exon does, use AUMsplice sdASO to specifically exclude that exon from the mRNA pool. The effect on the cell is then assessed. No knockout model is made, which would require deleting the exon from the genome, and the effect is reversible.

  • Protein engineering

    In some cases, skipping an exon can produce a desirable protein isoform. A researcher may intentionally skip an exon that encodes a domain to see how the truncated protein behaves. AUMsplice sdASO allows generation of such isoforms without genetic engineering.

  • Gene function rescue

    Exon skipping can sometimes rescue a gene's function by removing a problematic segment. For instance, if an exon contains a frame-shifting insertion, skipping that exon via AUMsplice sdASO may restore the downstream reading frame and partially rescue expression.

Every compatible cell type, 3D model, organoid and in vivo route, with the in vitro workflow.

See where it works

Experimental considerations

The working concentration is higher than that of a transfected oligonucleotide. The recommended working range for AUMsplice sdASOs is 5-20 μM. For steric blocking applications, higher concentrations may be required compared to RNA degradation approaches.

AUMsplice sdASOs redirect splicing rather than recruiting RNase H, so the readout is the ratio of splice variants rather than the total RNA. Total RNA may still fall where the new variant carries a premature termination codon, since the cell degrades such transcripts.

At the bench

Protocol, controls and what arrives

Add to culture medium or inject in vivo. The protocol, the controls and the How much to order page are written for this product.

View the protocolControls and validationHow much to orderHow to cite
Lead time
10-14 business days
Shipping
Products are shipped lyophilized, with priority shipping within the United States and international priority shipping elsewhere.
Quality control
Every oligo is strictly controlled for quality and is manufactured in an ISO 9001:2015 certified facility.

Related

Request a quote for AUMsplice sdASO

Provide us with your gene and exon of interest, and AUM designs the sequences for your target. A scientist is available before ordering to discuss the target and the cell type.

Request a quoteHow to citeView protocols

For research use only. Not for use in diagnostic or therapeutic procedures.