Transcript 5` Primer Design Worksheet

Molecular & Cellular Biology Lab
Instructor: Elmar Schmid, Ph.D.
(Student Name)
Primer Design Worksheet
A
B
5’ – T G C G T T A G G C G C G A A T T G T C A T G G T T A C G C C G T A A G G C T A T A G – 3’
C
D
3’ – A C G C A A T C C G C G C T T A A C A G T A C C A A T G C G G C A T T C C G A T A T C – 5’
1.
2.
3.
4.
5.
Write the nucleotide sequence of one suitable forward and reverse primer pair into
the empty white boxes below the selected target sequences A, B C or D (grey boxes)
Indicate the 5’ and 3’ ends of your selected primer pair
Draw arrows to indicate the polymerization direction of the Taq polymerase after
annealing of your chosen primer pair to the template DNA
What would be the length (in bp) of the PCR amplified DNA using your selected
pair above?
________ bp
What would be the calculated melting temperatures Tm of the two chosen (forward and
reverse) primers? (see “PCR” lab introduction section on my website for help)
Tm (forward) = ___ oC
6.
Tm (reverse) = ___ oC
Which annealing temperature TA would you chose to do your PCR with the two primers above?
(See “PCR” lab introduction section on my website for help)
TA = ___ oC