Comparison of embryonic human liver miRNA profiling studies with the chick profi

Question

Comparison of embryonic human liver miRNA profiling studies with the chick profiles found that several miRNAs appear to be expressed differently between the two species. For example, both miR-100 and miR-130b seem to be expressed higher in the embryonic avian liver than in the embryonic human liver. How would you design an experiment to determine which pathways these miRNAs regulate in the chick liver? Be sure to include an explanation of how each component of your experiment will aid you in identifying these pathways. You may use the luciferase assay, but you also need to include additional experiments to provide a more complete analysis of these miRNAs.

Explanation / Answer

Step 1: biotinylate your miRNA and one it in appropriate vectors. This will help us in subsequent steps to isolate miRNA-RNA complexes.

Step2- multiply/ enrich these miRNAs by transforming them in high copy number host cells.

Step3- isolate ur miRNA and allow them to tranfect a culture of embryonic chick liver cells.

Step4- after appropriate incubation time, obtain the cell lysate and allow it to run on streptavidin coated beads. This would cause biotin bound to the miRNAs to bind to them leading to isolation of miRNA-RNA complexes.

Step 5- elute them and go for RNA sequencing. This will help you identify the mRNA against/with whom the concerned miRNAs bind.

Step6- analysis of the target mRNAs would hep one deduce the pathway in which the miRNA be involved in.

Related Questions

Navigate

• Browse (All)
• Browse C
• Subjects

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.