Introduction:Here are is a
common PCR reaction. You should think of possible controls (both positive and negative)
you can perform along with the test reaction. Make a table. You should also consider
varying some of the components (DNA template concentration, Mg++ concentration,
etc) to see what works best. Be inventive. After you have performed the PCR and controls
you should analyze your results by gel electrophoresis on a 1% agarose gel.
We are going to use
the plasmid, C71794,
donated by Japanese organization, RGP as the DNA template for the PCR
reaction. The C71794 is partial cDNA of rice phytoene synthase gene
and was inserted between NotI and SacI at vector pBluescript
SK(-) II. The sequence of insert is available at Genbank (accession number
is C71794).
The primers are oligonucleotides that were designed by Dr. Wurtzel's lab
(#480(T3) and #481(T7)). There sequences are as follows: 480 - 5'
AATTAACCCTCACTAAAGGGA 3'; 481 - 5'
GTAATACGACTCACTATAGGGC 3'. T7 and T3
are very commonly primers at vector and used for sequencing. Here we can
use them to amplify the insert and detect its length .
Materials and recipes 
