PCR Activity - Answers
What 4 nucelotide primer would you use?
CGCA (complement to GCGT)
Once the primer binds, what sequence would add to the primer? (Remember, A pairs with T, C pairs with G)
CGCA(ACGCTAGCATTCAGC)TTACGTACGCCAGT (sequence doesn’t stop with the target sequence Taq polymerase keeps adding bases until the end of the strand)
The new strand is read right to left. What primer would you use to copy the sequence from the new strand of DNA?
GTAA (complement to CATT) - four nucleotides to the right of the target sequence, read backwards)
What sequence do you get after the nucleotides add to the second primer? Is this a copy of your target sequence and primers only? Is there any extra DNA included?
GTAA(GCTGAATGCTAGCGT)ACGC
Copy of target sequence and primers
No extra DNA in this strand
Why do you need to do more than one cycle of PCR in order to obtain just the target sequence?
Copy of target sequence and primers
No extra DNA in this strand
Bases add after the primer. The first primer binding and copying gets rid of all the DNA to the left of the target sequence. The second primer binding and copying gets rid of all the DNA to the right of the target sequence.
Why can’t you get a target sequence without primers on it?
Primers are always needed to start building more DNA
Once the PCR is done, you will have many different length strands of DNA. The smallest will be your target sequence. How can you separate out the target sequence DNA from the rest of the DNA?
The DNA can be separated in gel electrophoresis. Since the target sequence is the smallest, it will form a band at the bottom of the gel.