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<p>The <strong>SHAPE</strong> acronym stands for "<strong>S</strong>elective 2′ <strong>h</strong>ydroxyl <strong>a</strong>cylation analyzed by <strong>p</strong>rimer <strong>e</strong>xtension" | <p>The <strong>SHAPE</strong> acronym stands for "<strong>S</strong>elective 2′ <strong>h</strong>ydroxyl <strong>a</strong>cylation analyzed by <strong>p</strong>rimer <strong>e</strong>xtension". It is the primary method used to evaluate designs in [[EteRNA]] [[lab]] experiments.</p> | ||
<p>== Overview ==</p> | <p>== Overview ==</p> | ||
<p>== Background: The high-level structure of RNA ==</p> | |||
<p>== Background: Laboratory procedures ==</p> | |||
<p>=== PCR ===</p> | |||
<p>=== Transcription (DNA to RNA) ===</p> | |||
<p>=== Reverse transcription (RNA to DNA ) ===</p> | |||
<p>== A more detailed explanation of the SHAPE process ==</p> | |||
<p>== To be incorprated ==</p> | |||
<p>[[File:SHAPE reaction.png|thumb|400px]]This chemical probing strategy assesses local flexibility in [[RNA]] via accessibility of the [[ribose]] 2′-OH group to acylation by the electrophilic reagent NMIA or 1M7. In flexible regions (such as [[loop]]s, [[bulge]]s, and [[junction]]s), RNA adopts conformations that will promote formation of a nucleophilic 2′-oxyanion which reacts with NMIA or 1M7 to form a bulky 2′-O-adduct. Modified RNAs are subsequently evaluated by primer extension with an RNase H-deficient reverse transcriptase, creating a cDNA library corresponding to stops at sites of adduct formation in the RNA.</p> | <p>[[File:SHAPE reaction.png|thumb|400px]]This chemical probing strategy assesses local flexibility in [[RNA]] via accessibility of the [[ribose]] 2′-OH group to acylation by the electrophilic reagent NMIA or 1M7. In flexible regions (such as [[loop]]s, [[bulge]]s, and [[junction]]s), RNA adopts conformations that will promote formation of a nucleophilic 2′-oxyanion which reacts with NMIA or 1M7 to form a bulky 2′-O-adduct. Modified RNAs are subsequently evaluated by primer extension with an RNase H-deficient reverse transcriptase, creating a cDNA library corresponding to stops at sites of adduct formation in the RNA.</p> | ||
<p> </p> | <p> </p> | ||
Revision as of 19:14, 2 September 2013
The SHAPE acronym stands for "Selective 2′ hydroxyl acylation analyzed by primer extension". It is the primary method used to evaluate designs in EteRNA lab experiments.
== Overview ==
== Background: The high-level structure of RNA ==
== Background: Laboratory procedures ==
=== PCR ===
=== Transcription (DNA to RNA) ===
=== Reverse transcription (RNA to DNA ) ===
== A more detailed explanation of the SHAPE process ==
== To be incorprated ==
This chemical probing strategy assesses local flexibility in RNA via accessibility of the ribose 2′-OH group to acylation by the electrophilic reagent NMIA or 1M7. In flexible regions (such as loops, bulges, and junctions), RNA adopts conformations that will promote formation of a nucleophilic 2′-oxyanion which reacts with NMIA or 1M7 to form a bulky 2′-O-adduct. Modified RNAs are subsequently evaluated by primer extension with an RNase H-deficient reverse transcriptase, creating a cDNA library corresponding to stops at sites of adduct formation in the RNA.
== External links ==
- http://www.chem.unc.edu/rna/pdf-files/2005_em_jacs.pdf the original 2005 scientific paper
- The figure 1 of 'Probing Retroviral and Retrotransposon Genome Structures: The "SHAPE" of Things to Come.' presents a good overview of this method.
