A vector of 3000 bp has an EcoRI site at 400 and a HindIII site at 1000…
Contributed by StemPy LabTry it on paper first. A hint, the key idea and the worked solution are here when you need them.
A vector of bp has an EcoRI site at and a HindIII site at . An insert of bp is cloned into the EcoRI site. The insert has a HindIII site bp from its left end. The recombinant has bp. A student cuts the recombinant with HindIII and runs the digest.
- If the insert went in the same way as it is drawn, what are the two fragments?
- If it went in reversed, what are the two fragments?
- She finds bands of and bp. Which way did the insert go?
Need a reminder? The key ideaShow
Cut inside the cloning region of the vector, and not inside the gene. Two different enzymes give directional cloning and stop the vector from closing on itself. Recombinant vector stuffer insert. A diagnostic digest with an off-centre site shows the direction of an insert.
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. Vector sites: EcoRI , HindIII . Insert , with HindIII at from the left end.
Plan. Find the positions of the two HindIII sites in the recombinant. The vector HindIII site is at .
Step 1. (a) Same way: the insert site is at . The fragments are and bp.
Step 2. (b) Reversed: the insert site is at . The fragments are and bp.
Step 3. (c) Bands of and bp fit the reversed insert.
Answer: (a) and bp. (b) and bp. (c) Reversed.
Check. Both pairs add up to bp.
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