A PCR uses the primer 5′-GATCCGTTAGCAGTCCGA-3′…
Contributed by StemPy LabTry it on paper first. A hint, the key idea and the worked solution are here when you need them.
A PCR uses the primer 5′-GATCCGTTAGCAGTCCGA-3′.
- How many G and C bases does it have?
- What is its melting temperature by the Wallace rule?
- What annealing temperature would you try, if you use a temperature 5 °C below the melting temperature?
- A second primer has the same length but more A and T. Would you anneal it at a higher or a lower temperature?
Need a reminder? The key ideaShow
In a gel, the size of a fragment falls in a straight line against the distance on a log scale. A ring cut at sites gives pieces, and a straight DNA gives . In PCR, copies are made from one molecule in cycles. The Wallace rule is in degrees Celsius.
The worked solution
Have you tried it?
Work it out on paper first. It is fine to get stuck or make a mistake: that is how we learn. When you are ready, go through the solution one step at a time, and check each step against your own.
Given. The primer has 18 bases.
Plan. Count, then use the rule.
Step 1. (a) G and C: . A and T: .
Step 2. (b) .
Step 3. (c) .
Step 4. (d) A primer with more A and T has fewer hydrogen bonds, so a lower melting temperature. You would anneal it at a lower temperature.
Answer: ; ; ; lower.
Check. A primer of 18 bases with half G and C would melt at , close to the answer.
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