A primer is a short strand of RNA or DNA (generally about 18-22 bases) with a 3'-OH end that serves as a starting point for DNA synthesis. They identify sequences on the template DNA and upon providing required temperature, they anneal and start copying the sequence of the parent strand to form a new strand. Every primer has it's own annealing temperature which is usually 1-2 ⁰C lower than the melting temperature (Tm) of the primer.
- Primer Melting Temperature (Tm) by definition is the temperature at which one-half of the DNA duplex will dissociate to become single stranded and indicates the duplex stability.
Primers that are already known to amplify sequences in wheat are used in the parents and the mapping population of the current study.
- These primers come in vials lyophilized.
- Then they are diluted to concentration 100pMol/micro-litre. (stock) The volume of dilution is mentioned on the vials.
- Then working solution of primers is made by taking 100 microlitre of the primer stock and making up volume to 500 microlitres.
These primers are further used for screening the parents P1 and P2.
If the primers show polymorphism for the pair of parents, then they are proceeded for genotyping in the progeny.
