Haynes Lab:Notebook/Engineering PC-TFs/2012/11/26

Current revision

Engineering PC-TFs

Digest the pSB1A2/3 plasmid (from Rene) with XbaI & SpeI to get rid of the insert

Run entire 30 μL on a 1% gel (use big tooth comb) next to a ladder
You should see two bands. The larger band (2000 bp) is the backbone.
Cut out and gel purify this fragment (ignore the shorter fragment).
Notify Dr. Haynes when you are done. The next step is Gibson Assembly and transformation.

@@ Line 7: / Line 7: @@
 <!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
 ==Summary==
-*
+*'''[[User:Karmella Haynes|---Karmella]] 01:17, 26 November 2012 (EST)''':
+* Digest the pSB1A2/3 plasmid (from Rene) with XbaI & SpeI to get rid of the insert
+{| class="wikitable" border="0" cellspacing="3" <!-- Digest rxn. table -->
+|-valign="top"
+| <u>Reagent</u> || <u>Volume</u> || &nbsp;
+| rowspan="9" |
+|-
+| DNA (plasmid) || 20.0
+|-
+| 10x buffer || 3.0
+|-
+| XbaI || 1.0
+|-
+| SpeI || 1.0
+|-
+| dH<sub>2</sub>O || 5.0
+|-
+| &nbsp; || 30 μL --> 37°C/ ~10 min.
+|}
+* Run entire 30 μL on a 1% gel (use big tooth comb) next to a ladder
+* You should see two bands. The larger band (2000 bp) is the backbone.
+* Cut out and gel purify this fragment (ignore the shorter fragment).
+* Notify Dr. Haynes when you are done. The next step is Gibson Assembly and transformation.