|
Specificity of primers is very important for the generation of longer DNA amplification products. For example, if it is highly specific, even 20-mer primers can generate DNA fragments in length of greater than 20 kbp.
If possible, prepare primers according to the following criteria.
| a) | | The primer length should be 30 to 35 bases. |
| b) | The difference between the optimum annealing temperatures of paired primers should be within 2 to 3°C. |
| c) | Choose primers whose composition of G+C is around 50 to 60%. |
| d) | Avoid primer sequences which form hairpin loops or primer-dimers, especially at the 3'-end. |
| e) | Primers including inosin within its sequence sholud be avoided. |
|
