Rank the key steps necessary to select a ribozyme with ligase activity through S
ID: 167553 • Letter: R
Question
Rank the key steps necessary to select a ribozyme with ligase activity through Systematic Evolution of Ligands by Exponential Enrichment (SELEX) in order from first to last. Assume that you will complete multiple rounds of selection and that you want to diversify the pool each round. You add an RNA molecule that includes a sequence tag to the mixture of your potential ribozymes. If a ribozyme ligates the tag to its own 5 end, the ligating ribozyme can be purified out from the mix.
Rank the steps from first to last.
- Take a randomized RNA pool of a specific length.
- Transcribe back to RNA.
- Mix with RNA including the tag sequence.
- Select tagged RNAs and purify the ones larger than the original RNA including the tag sequence (indicates ligation).
- Amplify the ribozyme region with an error-prone DNA polymerase.
- Reverse-transcribe the selected RNAs to DNA.
Explanation / Answer
The steps from first to last will be
- Amplify the ribozyme region with an error prone DNA polymerase
- Take a randomized RNA pool of a specific length
- Transcribe back to RNA
- Mix with RNA including the tag sequence
- Select tagged RNAs and purify the ones larger than the original RNA including the tag sequence (indicates ligation)
- Reverse- transcribe the selected RNAs to DNA
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