Difference between revisions of "Part:BBa K2429168"

Latest revision as of 22:11, 25 October 2017

REST Guide Family Member

Parts K2429168 through K2429172 and parts K2429173 through K2429178 are a family of guide sequences used by either a dCas13a variant or is just a lone guide sequence to target the alternative exon in the mRNA that leads to the production of REST protein in cells. These guides lie downstream of a U6 promoter, which is a constitutive promoter typically used for the production of guide RNA. REST is a transcription factor that prohibits the transcription of oncogenes. When the mature mRNA for REST includes an alternative exon containing a stop codon, the resulting protein (known as REST4) is unable to prevent the transcription of oncogenes and can contribute to tumor formation. Application of our system in cells displaying the incorrect REST4 isoform attempts to correct this splicing error. Our team used multiple guide sequences to "tile" along the pre-mRNA to determine what part of the intron is best to target when trying to control its splicing efficiency. Based on the level of fluorescence associated with each guide, we can compare the results to see whether a specific targeted sequence is integral in the process of splicing, and whether the binding of the RNA binding protein to said sequence changes the splicing efficiency.

The guides shift by a few base pairs so we can make more precise conclusions about what intronic sequences are important for splicing.

The parts in the figure below are the actual sequences that lead to RNA guides that serve as a guide for the dCas13a protein. The sequence information will include show which specific sequence is downstream of the hU6 promoter.

The parts in the figure below are the actual sequences that lead to the production of antisense oligonucleotides (ASOs). The sequence information will include show which specific sequence is downstream of the hU6 promoter.

Sequence and Features