Description

Actinomycin D (CAS 50-76-0), also known as Dactinomycin, is an ultra-pure polypeptide antibiotic and a widely used transcription inhibitor. It functions as a DNA intercalator, forming a stable complex with double-stranded DNA and selectively inhibiting DNA-dependent RNA synthesis.

Because it binds DNA rather than RNA, Actinomycin D is frequently used in transcription shut-off experiments, mRNA half-life assays, RNA stability studies, and strand-specific reverse transcription workflows.

Mechanism of Action

Actinomycin D intercalates into double-stranded DNA and blocks RNA polymerase progression, thereby inhibiting DNA-primed RNA synthesis. It has also been reported to interfere with the minus-strand transfer step in reverse transcriptase systems.

Importantly, since Actinomycin D binds DNA but not RNA:

• It inhibits transcription from DNA templates

• It does not directly block RNA-templated first-strand cDNA synthesis

• It can suppress unwanted DNA-templated artifacts during RT-based workflows

This property makes it useful in directional RNA library preparation and strand-specific RT protocols.

Chemical structure: peptide

Specifications

Application

• Transcription inhibition studies

• mRNA stability and half-life assays

• Directional RNA library preparation

• HIV replication suppression research

• PC12 programmed cell death studies

• Selection agent in cell culture

Actinomycin D for mRNA Half-Life Assays (qPCR-Based Time Course)

One of the most searched use cases is transcription inhibition for mRNA decay studies.

Common Experimental Setup 

Researchers commonly report:

• Concentration range: 0.1–5 µg/mL (cell-type dependent)

• Typical time course: 2–24 hours

• Sampling intervals: every 2 hours for unstable RNAs

• Housekeeping control: 18S rRNA often used

However, optimal concentration depends on:

• Cell type (e.g., embryonic stem cells vs differentiated cells)

• Target RNA stability

• Cytotoxic sensitivity

Practical Recommendations

• Always optimize concentration for your cell line

• Start with 0.5–2 µg/mL for mammalian cells

• Use multiple housekeeping genes if RNA yield decreases significantly

• Confirm inhibition efficiency at early timepoints

Handling & Storage Considerations (Frequently Asked Questions)

Is Actinomycin D light sensitive?

Yes. It is highly light-sensitive. Always wrap containers in foil.

Does it stick to plastic or glass?

Yes. In solution, it tends to adsorb to surfaces and stain materials.

Should I prepare fresh stock?

Yes. Fresh DMSO stocks are recommended.

Is it stable?

• Stable 15+ months at 2–8°C if protected from light and moisture

• Diluted solutions degrade rapidly

• Frozen concentrated aliquots stable ~1 month

Documents

COA

SDS

Disclaimer: For laboratory research use only.