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The Importance of Primer Length
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Why are primers often between 15 and 20 base pairs long? Because at this length, the probability that the primer sequence is complementary to more than one sequence on the template DNA is very small. For example, what if your primer was only 1 base long? Then statistically, you'd expect that it would bind to 1 out of every 4 bases on the template DNA (If it is an "A", then it would bind to any "T".). If it were 2 bases long, then the probability that it would bind to any other sequence of two bases on the template DNA would be (1/4)x(1/4), or 1/16. The human genome is approximately 2 billion (2 x 10^9) bases long; if your template DNA is human genomic DNA, then you'd expect your primer to bind to (2 x 10^9) x (1/16) or 125 million different sites on the template! That's not very specific. However, what if your primer was 20 base pairs long? Now the probability of there being a sequence on the template DNA that was exactly complementary to the primer sequence is (2 x 10^9) x (1/4)^20, or about 1 in 5,000.
Strictly speaking, this statistical argument is only correct if we're talking about random DNA sequences. Since our genomes are by no means random, even primers of 20 base pairs in length and above are subject to a BLAST search to ensure that there are no other exact matches, other than your region of interest. |
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