page_title: DNA-directed RNA interference
section_title: Organization of ddRNAi constructs
hierarchical_section_title: DNA-directed RNA interference / Organization of ddRNAi constructs
caption_attribution_description: English: Aligned ddRNAi construct designs for the production of therapeutic shRNAs
context_page_description: DNA-directed RNA interference is a gene-silencing technique that utilizes DNA constructs to activate an animal cell's endogenous RNA interference pathways. DNA constructs are designed to express self-complementary double-stranded RNAs, typically short-hairpin RNAs, that once processed bring about silencing of a target gene or genes. Any RNA, including endogenous mRNAs or viral RNAs, can be silenced by designing constructs to express double-stranded RNA complementary to the desired mRNA target.
  This mechanism has great potential as a novel therapeutic to silence disease-causing genes. Proof-of-concept has been demonstrated across a range of disease models, including viral diseases such as HIV, hepatitis B or hepatitis C, or diseases associated with altered expression of endogenous genes such as drug-resistant lung cancer, neuropathic pain, advanced cancer and retinitis pigmentosa.
context_section_description: Figure 2 illustrates the most common type of ddRNAi DNA construct, which is designed to express a shRNA. This consists of a promoter sequence, driving expression of sense and antisense sequences separated by a loop sequence, followed by a transcriptional terminator. The antisense species processed from the shRNA can bind to the target RNA and specify its degradation. shRNA constructs typically encode sense and antisense sequences of 20 – 30 nucleotides. Flexibility in construct design is possible: for example, the positions of sense and antisense sequences can be reversed, and other modifications and additions can alter intracellular shRNA processing. Moreover, a variety of promoter loop and terminator sequences can be used.
  A particularly useful variant is a multi-cassette (Figure 2b). Designed to express two or more shRNAs, they can target multiple sequences for degradation simultaneously. This is a particularly useful strategy for targeting viruses. Natural sequence variations can render a single shRNA-target site unrecognizable preventing RNA degradation. Multi-cassette constructs that target multiple sites within the same viral RNA circumvent this issue.
caption: Figure 2: ddRNAi DNA constructs
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