User:Jonathan Dewerd/Notebook/TeamAllergy/2010/06/16

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Gene Silencing Vector

Error fetching PMID 11576441:
  1. Error fetching PMID 11576441: [Wesley-TPJ-2001]

Deals with the relative efficacy of various post-translational gene silencing (PTGS) techniques. Summary:

  • co-suppression (sRNA) Simply over-express a certain RNA and the cell's regulatory mechanisms will reduce the expression. 30% effective at suppressing GUS.
  • Adj-hp RNA (Adjacent to HairPin RNA): place the target RNA sequence next to a random hairpin and attach a single strand of the target RNA to one of the tails of the hairpin. 25% effective at suppressing GUS
  • hpRNA (HairPin RNA): let one of the hairpin tails be the target sequence. 55% effective.
  • ihpRNA (Intron on HairPin RNA): put part of an intron in the hairpin loop. 90% effective. Overhang makes it slightly less effective.

GUS is a standard reporting gene. When activated, things turn blue.

PCR

The primers arrived today so we were able to PCR the cDNA we made yesterday. We used the Fraa1_{Antisense,Sense}_{FR} promoters. "Sense" and "Antisense" refer to the final product: Sense_F and Sense_R go together. We used the protocol on the protocol page with the Phusion Polymerase. We also poured the gel which we'll run tomorrow.


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