Difference between revisions of "20.109(F07):Module 2"
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==Module 2== | ==Module 2== | ||
| − | '''Instructors:''' [ | + | '''Instructors:''' [http://openwetware.org/wiki/Natalie_Kuldell ] and [http://openwetware.org/wiki/User:AgiStachowiak | Agi Stachowiak] |
| − | '''TA:''' | + | '''TA:'''[http://openwetware.org/wiki/User:AliceLo | Alice Lo] |
| − | In this experiment, we will consider unintended and unpredicted effects of an experimental perturbation. Our goal is a precise one, namely to silence gene expression of a measurable gene, luciferase, using RNA interference (RNAi). Each group will begin by designing a short interfering RNA (siRNA) against luciferase, but as we'll see, siRNAs can vary in efficacy and specificity. After transfecting a mammalian cell line with the siRNA | + | In this experiment, we will consider unintended and unpredicted effects of an experimental perturbation. Our goal is a precise one, namely to silence gene expression of a measurable gene, luciferase, using RNA interference (RNAi). Each group will begin by designing a short interfering RNA (siRNA) against luciferase, but as we'll see, siRNAs can vary in efficacy and specificity. After transfecting a mammalian cell line with the siRNA you ve designed and a reporter plasmid, we will evaluate the silencing using a luciferase assay and microarray technology. The first assay evaluates the efficacy of the siRNA in silencing. The second assay gives genome-wide expression data to reveal the specificity of your siRNA for the gene you ve targeted. Through this combined approach, we'll assess the balance of targeted and off-target effects. |
[[Image:Macintosh_HD-Users-nkuldell-Desktop-sprinkler.jpg|400px|center]] | [[Image:Macintosh_HD-Users-nkuldell-Desktop-sprinkler.jpg|400px|center]] | ||
| − | [[20.109(F07): | + | [[20.109(F07): siRNA design | Module 2 Day 1: siRNA design and introduction to cell culture]]<br> |
| − | [[20.109(F07): | + | [[20.109(F07): Transfection | Module 2 Day 2: Transfection]]<br> |
| + | [[20.109(F07): Luciferase assays and RNA prep| Module 2 Day 3: Luciferase assays and RNA prep]]<br> | ||
| + | [[20.109(F07): Journal article discussion| Module 2 Day 4: Journal article discussion]]<br> | ||
| + | [[20.109(F07): cDNA synthesis and microarray| Module 2 Day 5: cDNA synthesis and microarray]]<br> | ||
| + | [[20.109(F07): Microarray data analysis| Module 2 Day 6: Microarray data analysis]]<br> | ||
| + | [[20.109(F07): Mod 2 Day 7| Module 2 Day 7: Reports due]]<br> | ||
| + | [[20.109(F07): Module 2 oral presentations| Module 2 Day 8: Oral Presentations]]<br> | ||
| + | direct link to [[20.109(F07): Expression engineering report| writing assignment]]<br> | ||
| + | |||
| + | [http://openwetware.org/wiki/20.109(F07):_TA's_notes_for_module_2 | TA notes, mod 2] | ||
Latest revision as of 15:54, 15 June 2015
Module 2
Instructors: [1] and | Agi Stachowiak
TA:| Alice Lo
In this experiment, we will consider unintended and unpredicted effects of an experimental perturbation. Our goal is a precise one, namely to silence gene expression of a measurable gene, luciferase, using RNA interference (RNAi). Each group will begin by designing a short interfering RNA (siRNA) against luciferase, but as we'll see, siRNAs can vary in efficacy and specificity. After transfecting a mammalian cell line with the siRNA you���ve designed and a reporter plasmid, we will evaluate the silencing using a luciferase assay and microarray technology. The first assay evaluates the efficacy of the siRNA in silencing. The second assay gives genome-wide expression data to reveal the specificity of your siRNA for the gene you���ve targeted. Through this combined approach, we'll assess the balance of targeted and off-target effects.
Module 2 Day 1: siRNA design and introduction to cell culture
Module 2 Day 2: Transfection
Module 2 Day 3: Luciferase assays and RNA prep
Module 2 Day 4: Journal article discussion
Module 2 Day 5: cDNA synthesis and microarray
Module 2 Day 6: Microarray data analysis
Module 2 Day 7: Reports due
Module 2 Day 8: Oral Presentations
direct link to writing assignment
