Team:SYSU-Software/Models2

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         <h1 id="doctitle" class="pagetitle"><span lang="EN-US">Algorithm——iRNA
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         <h1 id="doctitle" class="pagetitle" style="line-height: 1.5em;"><span lang="EN-US">Algorithm——</span>S<span lang="EN-US">iRNA
designer</span>&amp; <span lang="EN-US">Riboswitch designer</span></h1>
designer</span>&amp; <span lang="EN-US">Riboswitch designer</span></h1>
<h2><span lang="EN-US">siRNA designer:<o:p></o:p></span></h2>
<h2><span lang="EN-US">siRNA designer:<o:p></o:p></span></h2>
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w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;per AU end&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:e&gt;&lt;/m:d&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;+&lt;/m:t&gt;&lt;/m:r&gt;&lt;m:sSub&gt;&lt;m:sSubPr&gt;&lt;m:ctrlPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;/m:ctrlPr&gt;&lt;/m:sSubPr&gt;&lt;m:e&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot; w:hint=&quot;fareast&quot;/&gt;&lt;wx:font wx:val=&quot;瀹嬩綋&quot;/&gt;&lt;th&quot;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;螖&lt;/m:t&gt;&lt;/m:r&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;G掳h&quot;37&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:e&gt;&lt;m:sub&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;37 symmetry&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:sub&gt;&lt;/m:sSub&gt;&lt;m:d&gt;&lt;m:dPr&gt;&lt;m:ctrlPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;/m:ctrlPr&gt;&lt;/m:dPr&gt;&lt;m:e&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;self-complementary duplexes&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:e&gt;&lt;/m:d&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;+&lt;/m:t&gt;&lt;/m:r&gt;&lt;m:nary&gt;&lt;m:naryPr&gt;&lt;m:chr m:val=&quot;鈭?/&gt;&lt;m:limLoc m:val=&quot;undOvr&quot;/&gt;&lt;m:subHide m:val=&quot;1&quot;/&gt;&lt;m:supHide m:val=&quot;1&quot;/&gt;&lt;m:ctrlPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:a-ascii=&quot;Cambria Math&quot; 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w:h-ansi=&quot;Cambria Math&quot; w:hint=&quot;fareast&quot;/&gt;&lt;wx:font wx:val=&quot;瀹嬩綋&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;?&quot;/&gt;?/m:t&gt;&lt;/m:r&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;G掳37&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:e&gt;&lt;m:sub&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=/&gt;&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt; stacking&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:sub&gt;&lt;/m:sSub&gt;&lt;m:r&gt;&lt;m:rPr&gt;&lt;m:sty m:val=&quot;p&quot;/&gt;&lt;/m:rPr&gt;&lt;w:rPr&gt;&lt;w:rFonts w:ascii=&quot;Cambria Math&quot; w:h-ansi=&quot;Cambria Math&quot;/&gt;&lt;wx:font wx:val=&quot;Cambria Math&quot;/&gt;&lt;w:sz w:val=&quot;28&quot;/&gt;&lt;w:sz-cs w:val=&quot;28&quot;/&gt;&lt;/w:rPr&gt;&lt;m:t&gt;]&lt;/m:t&gt;&lt;/m:r&gt;&lt;/m:e&gt;&lt;/m:nary&gt;&lt;/m:oMath&gt;&lt;/m:oMathPara&gt;&lt;/w:p&gt;&lt;w:sectPr wsp:rsidR=&quot;00000000&quot; wsp:rsidRPr=&quot;006447FA&quot;&gt;&lt;w:pgSz w:w=&quot;12240&quot; w:h=&quot;15840&quot;/&gt;&lt;w:pgMar w:top=&quot;1440&quot; w:right=&quot;1800&quot; w:bottom=&quot;1440&quot; w:left=&quot;1800&quot; w:header=&quot;720&quot; w:footer=&quot;720&quot; w:gutter=&quot;0&quot;/&gt;&lt;w:cols w:space=&quot;720&quot;/&gt;&lt;/w:sectPr&gt;&lt;/wx:sect&gt;&lt;/w:body&gt;&lt;/w:wordDocument&gt;">
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<ul>
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<a href="https://2012.igem.org/Team:SYSU-Software/">Main Page</a>
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<a href="https://2012.igem.org/Team:SYSU-Software">Main Page</a>
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<a href="https://2012.igem.org/Team:SYSU-Software/Accomplishment">Accomplishment</a>
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<a href="https://2012.igem.org/Team:SYSU-Software/Download">Source Code</a>
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documentation-user-guide-main-reminderfox-window">Source Code</a>
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<a href="/documentation-user-guide-options-and-preferences">Algorithm-FBA</a>
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<a href="">Uninstalling</a>
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<a href="https://2012.igem.org/Team:SYSU-Software/Uninstalling">Uninstalling</a>
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Latest revision as of 07:16, 25 September 2012

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Algorithm——SiRNA designer& Riboswitch designer

siRNA designer:

Tom Tuschl's rule (for cDNA)

1. Select targeted region from a given cDNA sequence beginning 50-100 nt downstream of start codon

2. Look for sequence motif AA+N19+TT first. If there is no suitable sequence look 23-nt sequence motif NA+N21 and convert the 3' end of the sense siRNA to TT

3. Or search for NAR+N17+YNN

PS: Target sequence should have a GC content of around 50%

R means A/G, and Y means T/C.

Rational siRNA design(for mRNA)

Evaluate potential candidates and assign scores to them, sequences with higher scores will have higher chance of success in RNAi.

The table below lists the 8 criteria and the methods of score assignment.

Criteria

Description

Score

Yes

No

1

Moderate to low (30%-52%) GC Content

1 point

 

2

At least 3 A/Us at positions 15-19 (sense)

1 point /per A or U

 

3

Melting temperature-Tm*<20

1 point

 

4

A at position 19 (sense)

1 point

 

5

A at position 3 (sense)

1 point

 

6

U at position 10 (sense)

1 point

 

7

No G/C at position 19 (sense)

 

-1 point

8

No G at position 13 (sense)

 

-1 point

The "anti-sense" strand is the siRNA strand that is complementary to the target mRNA and that will be binding to the mRNA.

The melting temperature’ of a siRNA candidate can be calculated by the formula showed below:

https://static.igem.org/mediawiki/igem.org/e/e8/Image101.png

wA, xT, yG, zC are separately the number of A, T, G, C in a siRNA candidate.

All siRNA candidates scored higher than 6 are acceptable.


 

Riboswitch designer:

The designer is based on a special promoter working in eukaryotes - internal ribosome entry site (IRES). We design the upstream sequence that can bind to IRES part so the second structure of IRES can be transformed by the ligand.

Up regulated:

Target sequence is assembled by five parts: MS-SL, aaIRES (anti-anti-IRES), aptamer, aIRES (anti-IRES), IRES.

https://static.igem.org/mediawiki/igem.org/e/ec/Image002.png

The IRES, aIRES, aaIRES parts are settled, we get aptamer sequence according to customer’s quests from aptamer database and the MS-SL is paired to the combination part of aaIRES and aptamer. Then adjust the MS-SL part’s length to fit it’s free energy to -11.7 kcal/mol.

 

Down regulated:

The differences from down regulated to up regulated riboswitch are the order between aptamer and aaIRES, the sequence of MS part. The new structure is showed below:

https://static.igem.org/mediawiki/igem.org/9/90/Image003.jpg

We set MS to “CCUCU” under experimental data and the aptamer still comes from aptamer database.

 

Algorithm about free energy calculation comes from Turner’s team:

Hairpin loops:

https://static.igem.org/mediawiki/igem.org/8/89/Image004.png

In this equation, n (5) is the number of nucleotides in loop, the terminal mismatch parameter is the sequence-dependent term for the first mismatch stacking on the terminal base pair.

Watson-Crick Helices:

https://static.igem.org/mediawiki/igem.org/3/3c/Image005.png

Specific data can be found on web site:

http://rna.urmc.rochester.edu/NNDB/turner04/index.html

 

References:

1.Elbashir SM et al. (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature. 411:494-498.

2.Elbahir SM et al. (2001). Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J. 20:6877-6888.

3.Elbashir SM et al. (2002). Analysis of gene function in somatic mammalian cells using small interfering RNAs. Methods. 26:199-213.

4.Reynolds A, Leake D, Boese Q, Scaringe S, Marshall WS, Khvorova A. Rational siRNA design for RNA interference. Nat Biotechnol. 2004 Mar;22(3):326-30.

5.Ogawa, A. (2011). Rational design of artificial riboswitches based on ligand-dependent modulation of internal ribosome entry in wheat germ extract and their applications as label-free biosensors. RNA (New York, N.Y.), 17(3), 478-88. doi:10.1261/rna.2433111

6.Ogawa, A. (2012). Rational construction of eukaryotic OFF-riboswitches that downregulate internal ribosome entry site-mediated translation in response to their ligands. Bioorganic & medicinal chemistry letters, 22(4), 1639-42. Elsevier Ltd. doi:10.1016/j.bmcl.2011.12.118

7.Turner, D. H. & Mathews, D. H.  (2009).  NNDB: The nearest neighbor parameter database for predicting stability of nucleic acid secondary structure.  Nucleic Acids Research.  38, D280-D282.

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