Although employing RNAi vector systems can be slightly more involved than using synthetic RNAi reagents, the flexibility of the vector based systems is compelling for many RNAi researchers conducting both in vitro and in vivo experiments.

There are two main types of RNAi expression vector technologies on the market: short hairpin (shRNA) expression vectors and artificial microRNA (miRNA) expression vectors. Most RNAi vectors available today employ shRNA vector technology, which typically involves shRNA expression from a Pol III promoter and may or may not employ viral delivery.

These RNAi vectors express shRNA sequences, typically from a U6 or H1 promoter, and some have inducible promoters (typically H1/TO—a tetracycline-inducible promoter).

While these vectors can be used for in vivo RNAi experiments, there are some drawbacks, including low design success rate and inability to track shRNA expression or express the shRNA in a specific target tissue.

RNAi Vector Delivery Method
Similar to RNAi vectors for in vitro applications, you can use either standard transfection techniques or a viral delivery method to deliver RNAi vectors in vivo.

Currently, the delivery of an RNAi expression vector in vivo without using a viral RNAi delivery system is similar to delivering synthetic dsRNA in vivo.

Typically, this would involve complexing the RNAi expression vector with a commonly used lipid-based in vitro transfection reagent and injecting directly into the animal. While this may be the easiest approach for delivering RNAi vectors into animals, it has quite a few limitations, including the inability to deliver systemically, and low transfection efficiencies.

Viral RNAi Delivery Method
For these reasons, most researchers choose to use a viral delivery method when employing RNAi vectors for in vivo experiments. Regardless of whether one chooses an shRNA or miR RNAi vector system, viral delivery is a huge advantage for many in vivo approaches.

Most viral RNAi delivery approaches involve either adenoviral, retroviral (non-lentiviral), or lentiviral RNAi technologies:

  • Adenovirus can be used for transient RNAi expression in either dividing or non-dividing cells
  • Retrovirus can be employed for transient or stable expression, but can only be used to transduce dividing cells
  • Lentiviral delivery affords the most options, as it can be used for transient or stable expression in dividing or nondividing cells, as well as neuronal cells, drug- or growth-arrested cells, or even primary cells

To learn more about RNAi technologies watch this In Vitro to In Vivo webinar. In this informative webinar, Life Technologies scientist Sasha Vlassov presents our siRNA and miRNA technologies, namely Silencer Select siRNAs and mirVana Mimics and Inhibitors, and their respective application for mRNA and miRNA functional analysis in vitro and in vivo.

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