Haynes Lab:Notebook/Investigating Photo-Switchable Synthetic Nucleosomes/2012/11/06: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(Autocreate 2012/11/06 Entry for Haynes_Lab:Notebook/Investigating_Photo-Switchable_Synthetic_Nucleosomes) |
|||
(23 intermediate revisions by 2 users not shown) | |||
Line 6: | Line 6: | ||
| colspan="2"| | | colspan="2"| | ||
<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | <!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | ||
== | ==November 6, 2012== | ||
* | * Restriction digests of H2B-GFP and LOV | ||
{|border="1" cellpadding="5" cellspacing="0" align="center" | |||
|+Restriction Digest Reagents | |||
|- | |||
! scope="col" style="background:#efefef;" | Reagent | |||
! scope="col" style="background:#efefef;" | H2B-GFP/EcoRI | |||
! scope="col" style="background:#efefef;" | H2B-GFP/PstI | |||
! scope="col" style="background:#efefef;" | LOV/PstI | |||
|- | |||
|DNA(plasmid) | |||
|2.0 | |||
|2.0 | |||
|2.0 | |||
|- | |||
|10x buffer | |||
|1.5 | |||
|1.5 | |||
|1.5 | |||
|- | |||
|EcoRI | |||
|1.0 | |||
|0 | |||
|0 | |||
|- | |||
|PstI | |||
|0 | |||
|1.0 | |||
|1.0 | |||
|- | |||
|dH<sub>2</sub>O | |||
|10.5 | |||
|10.5 | |||
|10.5 | |||
|- | |||
|Total | |||
|15.0 | |||
|15.0 | |||
|15.0 | |||
|} | |||
{| class="wikitable" border=0 | |||
|- | |||
| <u>Expected:</u><br>1. H2B-GFP/ EcoRI = 4885, 210 <br>2. H2B-GFP/ PstI = 5101<br>3. LOV/PstI = 251 | |||
| <br> | |||
|} | |||
<center> | |||
[[Image:VN_Gel_11-6-12.png|150px|Results of the digests are shown.]] | |||
[[Image:KAH_Fermentas_GeneRuler_1kbplus.jpg]] | |||
</center> | |||
<center> | |||
Results of the digests are shown. | |||
</center> | |||
---- | |||
*'''[[User:Karmella Haynes|---Karmella]] 16:01, 7 November 2012 (EST)''': Note: Please re-do LOV/PstI | |||
<br> | |||
*'''[[User:Karmella Haynes|---Karmella]] 16:01, 7 November 2012 (EST)''': Here is a reaction table for PCR | |||
* First, resuspend dry primer oligos (in blue-capped tubes) to 100 μM | |||
* Then make working solutions of 10 μM | |||
* Reactions: | |||
# H2B-GFP plasmid + BB_H2B fwd + BB_H2B rev | |||
# LOV plasmid + BB_LOV fwd + BB_LOV rev | |||
{| class="wikitable" border=1 cellpadding="5" cellspacing="0" | |||
|- | |||
| Reagents || H2B || LOV | |||
|- | |||
| Plasmid DNA || 0.5 μL || 0.5 | |||
|- | |||
| primer 1 (10 μM) || 1.0 || 1.0 | |||
|- | |||
| primer 2 (10 μM) || 1.0 || 1.0 | |||
|- | |||
| 2x GoTaq mix || 12.5 || 12.5 | |||
|- | |||
| dH<sub>2</sub>O || 10.5 || 10.5 | |||
|} | |||
PCR program: | |||
* 95°C, 3 min. | |||
* [95°C, 30 sec; 57°C, 30 sec.; 72°C, 30 sec.] x35 cycles | |||
* 72°C, 3 min. | |||
* 4°C ∞ | |||
<!-- ##### DO NOT edit below this line unless you know what you are doing. ##### --> | <!-- ##### DO NOT edit below this line unless you know what you are doing. ##### --> |
Revision as of 02:52, 14 November 2012
Project name | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> | ||||||||||||||||||||||||||||||||||||||||||||||||
November 6, 2012
Results of the digests are shown.
PCR program:
|