Endoribonuclease-prepared siRNA

esiRNA or Endoribonuclease-prepared siRNAs are a mixture of siRNA oligos resulting from cleavage of long double-stranded RNA (dsRNA) with an endoribonuclease such as Escherichia coli RNase III or Dicer.[1][2]

An alternative concept to the usage of chemically synthesized siRNA for RNA Interference (RNAi) is the enzymatic digestion of long double stranded RNAs in vitro. In this case a cDNA template is amplified by PCR and tagged with two bacteriophage-promotor sequences. RNA polymerase is then used to generate long double stranded RNA that is homologous to the target-gene cDNA. This RNA is subsequently digested with RNase III from Escherichia coli to generate short overlapping fragments of siRNAs with a length between 18-25 base pairs. This complex mixture of short double stranded RNAs is similar to the mixture generated by Dicer cleavage in vivo and is therefore called endoribonuclease-prepared siRNA or short esiRNA. esiRNA are a heterogeneous mixture of siRNAs that all target the same mRNA sequence. These multiple silencing triggers lead to highly specific and effective gene silencing.[2]

References

  1. ↑ Blow, Nathan (2008). "RNAi technologies: a screen whose time has arrived". Nature Methods. 5: 361–368. doi:10.1038/nmeth0408-361. Retrieved 7 September 2017.
  2. 1 2 Kittler, R; Pelletier, L (2004). "An endoribonuclease-prepared siRNA screen in human cells identifies genes essential for cell division". Nature. 432: 1036–1040. doi:10.1038/nature03159.

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