1. Into a sterile 0.75ml tube add the following. Take care to use sterile tips and avoid cross contamination. Use new tips for each reagent and template.

Reagent	Concentration	Amount(µl)
25 mM MgCl2	1.5 mM	3
10 mM dNTPs	200 µM	1
10× buffer	1x	5
primer 1
(10-25 pmol/µl)	0.2-0.5 pmol/µl	1
primer 2
(10-25 pmol/µl)	0.2-0.5 pmol/µl	1
Taq DNA
polymerase (5U/µl)	2.5 U	0.5
Reverse Transcriptase
product (template)		5
DEPC water		33.5
Total volume:		50

2. Add 20 µl of mineral oil to the top of mixture, spin, and run in the thermocycler as follows:

3. Take 5 µl of PCR product and check it by examination on an agarose gel (PCR Products).