Desgined
Primers from contigs of Turmeric and Ginger
Primers
were designed using primer3 online tool. This
software tool will provide one forward and reverse
primer and four additional oligos. First two primers
are quality primers additional oligos are less
quality primers. By checking the GC content, melting
temperature, self complimentarity and mispriming
of primer will determined the quality of the primer.
Some
thoughts on designing primers.
1. primers should be 17-28
bases in length;
2. base composition should be 50-60% (G+C);
3. primers should end (3') in a G or C, or CG
or GC: this prevents "breathing" of
ends and increases efficiency of priming;
4. Tms between 55-80°C are preferred;
5. 3'-ends of primers should not be complementary
(ie. base pair), as otherwise primer dimers
will be synthesised preferentially to any other
product;
6. primer self-complementarity (ability to form
2° structures such as hairpins) should be
avoided;
7. runs of three or more Cs or Gs at the 3'-ends
of primers may promote mispriming at G or C-rich
sequences (because of stability of annealing),
and should be avoided.
These
designed primers were quality checked using
FAST PCR program and PCR product size
