SBB12Ntbk-GregFukushima: Difference between revisions
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==[[User:Greg_Fukushima|Greg Fukushima]] | ==[[User:Greg_Fukushima|Greg Fukushima]], 7 February 2012== | ||
This is my first day of notebooks for 140L! I love wikis! | This is my first day of notebooks for 140L! I love wikis! | ||
==[[User:Greg_Fukushima|Greg Fukushima]] | ==[[User:Greg_Fukushima|Greg Fukushima]], 16 February 2012== | ||
First Day of PCR and cloning: | |||
Protocol for PCR: | |||
The oligo concentrations in your stocks should be 100uM. You use them at 10uM in this protocol. So, you first need to make an oligo dilution of: | |||
9uL Water | |||
1uL 100uM oligo | |||
You can throw away the remainder of the diluted oligo when you are done, but hold onto your stock tube! | |||
Set up the following reaction in a PCR tube: | |||
<pre> | |||
24uL ddH2O | |||
3.3uL 10x Expand Buffer "2" | |||
3.3uL dNTPs (2mM in each) | |||
1uL Oligo 1, 10uM | |||
1uL Oligo 2, 10uM | |||
0.5uL Expand polymerase "1" | |||
0.5uL Template DNA | |||
</pre> | |||
Revision as of 20:50, 18 February 2012
Greg Fukushima, 7 February 2012
This is my first day of notebooks for 140L! I love wikis!
Greg Fukushima, 16 February 2012
First Day of PCR and cloning:
Protocol for PCR: The oligo concentrations in your stocks should be 100uM. You use them at 10uM in this protocol. So, you first need to make an oligo dilution of:
9uL Water 1uL 100uM oligo
You can throw away the remainder of the diluted oligo when you are done, but hold onto your stock tube!
Set up the following reaction in a PCR tube:
24uL ddH2O 3.3uL 10x Expand Buffer "2" 3.3uL dNTPs (2mM in each) 1uL Oligo 1, 10uM 1uL Oligo 2, 10uM 0.5uL Expand polymerase "1" 0.5uL Template DNA
