Daniel:Notebook/ComboLock/2016-12-9: Difference between revisions
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>Djacobse (Created page with "=Lock Oligo Protocol= Back to Calendar This protocol covers a new method using ideas taken from Weibrecht ''et al'' New Biotechnology 2012 (Lan...") |
>Djacobse |
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[[Image:C_Probes-Original.png|500px|C probes in the orientation of their original design. Template (mRNA/control oligo) is in black, C probes in red. Vertical line indicates 5' end, arrow indicates 3' end.]] | [[Image:C_Probes-Original.png|500px|C probes in the orientation of their original design. Template (mRNA/control oligo) is in black, C probes in red. Vertical line indicates 5' end, arrow indicates 3' end.]] | ||
When | When subjected to polymerization, if the latch is not present this yields a product that follows the backbone of the second C probe. See image below for clarification. | ||
[[Image:C_Probes-BadCircularization.png|500px|C probes gap-filled when padlock binds (but not latch). C probes in red, padlock probe in green and polymerase-added bases in magenta. Vertical line indicates 5' end, arrow indicates 3' end.]] | |||
==Protocol== | ==Protocol== |
Revision as of 18:31, 9 December 2016
Lock Oligo Protocol
This protocol covers a new method using ideas taken from Weibrecht et al New Biotechnology 2012 (Landegren lab). The main idea I see is that they almost never use polymerase, but prefer to instead use ligation events as the predominant form. Based on the results from the extended latch experiment, the RCA-based cell test, and the sequencing from the first circularization test, it seems the probes do not circularize correctly. Note the original orientation of the C probes with the 5' end being the genome matching region and the 3' end being the latch/padlock adapter.
When subjected to polymerization, if the latch is not present this yields a product that follows the backbone of the second C probe. See image below for clarification.