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Team:University College London/Protocols/PrimerDesign
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Primers should be designed to: | Primers should be designed to: | ||
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1) Be 18-25 nucleotides in length | 1) Be 18-25 nucleotides in length | ||
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2) Have a melting temperature that does not differ by more than 5oC between the two primers of a pair | 2) Have a melting temperature that does not differ by more than 5oC between the two primers of a pair | ||
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3) GC nucleotides should constitute 40-60% | 3) GC nucleotides should constitute 40-60% | ||
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4) Have a 3’primer end that is not complimentary to any region of the primer pair | 4) Have a 3’primer end that is not complimentary to any region of the primer pair | ||
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5) Contain an equal distribution of the nucleotides | 5) Contain an equal distribution of the nucleotides | ||
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6) Closely match the target sequence (and so known polymorphic sites are avoided) | 6) Closely match the target sequence (and so known polymorphic sites are avoided) | ||
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7) Have a GC clamp on either end | 7) Have a GC clamp on either end | ||
Latest revision as of 10:13, 22 August 2012
Primer Design
Primers should be designed to:
1) Be 18-25 nucleotides in length
2) Have a melting temperature that does not differ by more than 5oC between the two primers of a pair
3) GC nucleotides should constitute 40-60%
4) Have a 3’primer end that is not complimentary to any region of the primer pair
5) Contain an equal distribution of the nucleotides
6) Closely match the target sequence (and so known polymorphic sites are avoided)
7) Have a GC clamp on either end
