Select aptamers against nucleic acid-like targets

Projects Andy thought might yield results quickly.

1.) Design oligonucleotide sequences (pick length, composition, sequence).

  • How do these variables affect the answers you're likely to get?

2.) Order the same oligonucleotide as both DNA and PNA (peptide nucleic acid), with a biotin moiety.

  • What are PNAs, what do they look like, where would you get them from?

3.) Immobilize on streptavidin-coated beads.

  • What is the basis for streptavidin-biotin interactions?
  • Where would you get these beads from?

4.) Select RNA binding sequences based on a centrifugation partition.

  • What are possible artefacts?
  • What controls or simultaneous experiments could you run to prevent these?

5.) After multiple rounds, clone and sequence binding species. Compare binding motifs for DNA versus PNA.

  • How will you identify binding motifs?

6.) Determine how many binding motifs you obtained; compare with how many binding motifs you expected.

  • How will you calculate how many binding motifs were expected?

Further questions and issues:

  • Has anyone done this before? What does the primary literature look like?
  • PNAs can potentially be used as antisense drugs. How might your findings influence the design of these drugs?
  • If you used a library that could be introduced into real-time PCR reactions, you could actually quantify the progress of the selection. What is real-time PCR? How does it work? How can it be used to quantitate amplicons?
  • How could your data be used for calculating the delta G of binding of a given PNA base-pair?

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