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The melting Temperature Tm is calculated as followed (Kibbe, 2007): | The melting Temperature Tm is calculated as followed (Kibbe, 2007): | ||
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$$ T_m = 79.8 + 18.5 * log_10([Na^+]) + (58.4 * [G/C content])+(11.8*([G/C content])^2) - \left \frac{820}{length siRNA} \right$$ | $$ T_m = 79.8 + 18.5 * log_10([Na^+]) + (58.4 * [G/C content])+(11.8*([G/C content])^2) - \left \frac{820}{length siRNA} \right$$ | ||
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The tool checks each criterion and only considers siRNAs with a score higher than six for further steps. | The tool checks each criterion and only considers siRNAs with a score higher than six for further steps. | ||
</article> | </article> | ||
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The initial hypothesis is that the given siRNA effectively silences an mRNA. To perform the calculations a prior probability is necessary. The prior probability for effective gene silencing of mammalian genes can be obtained from former siRNA experiments and is approximately 0.1. Since we have no data on prokaryotic siRNAs, we use the same prior probability for our prediction. </br> | The initial hypothesis is that the given siRNA effectively silences an mRNA. To perform the calculations a prior probability is necessary. The prior probability for effective gene silencing of mammalian genes can be obtained from former siRNA experiments and is approximately 0.1. Since we have no data on prokaryotic siRNAs, we use the same prior probability for our prediction. </br> | ||
The gene silencing probability can be described as: | The gene silencing probability can be described as: | ||
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\(P^{eff}\) is the prior probability 0.1 as mentioned above. The siRNA sequence is represented by \(X\), where \(X_1, X_2 ... X_n\) belong to the possible nucleotides adenine, guanine, cytosine and thymine. As \(P(X|eff)\) is the probability, that the given siRNA sequence will effectively silence if the nucleotides belong to the frequent nucleotides of common effective siRNAs, it is computed as the product of the probabilities that a particular nucleotide is located at a particular position of the siRNA: | \(P^{eff}\) is the prior probability 0.1 as mentioned above. The siRNA sequence is represented by \(X\), where \(X_1, X_2 ... X_n\) belong to the possible nucleotides adenine, guanine, cytosine and thymine. As \(P(X|eff)\) is the probability, that the given siRNA sequence will effectively silence if the nucleotides belong to the frequent nucleotides of common effective siRNAs, it is computed as the product of the probabilities that a particular nucleotide is located at a particular position of the siRNA: | ||
$$ P(X|eff) = \prod_{i=1}^{19} q_{x_i^n}^{eff} \qquad (2)$$ | $$ P(X|eff) = \prod_{i=1}^{19} q_{x_i^n}^{eff} \qquad (2)$$ |
Revision as of 17:37, 14 October 2018
siRCon - A siRNA Constructor
siRNAS short introduction
Choosing appropriate design methods
Rational siRNA design
Rule | Score |
---|---|
30%-52% G/C content | +1 |
At least 3 'A/U' bases at positions 15-19 | +1 (for each 'A/U' base) |
Absence of internal repeats (\(T_m \lt 20\)) | +1 |
An 'A' base at position 3 | +1 |
An 'A' base at position 19 | +1 |
An 'U' base at position 19 | +1 |
A base other than 'G' or 'C' at 19 | -1 |
A base other than 'G' at position 13 | -1 |
Ui-Tei rule
- An ‘A’ or ‘T’ at position 19
- A ‘G’ or ‘C’ at position 1
- At least five ‘U’ or ‘A’ residues from positions 13 to 19
- No ‘GC’ stretch more than 9nt long