Team:Groningen/21 June 2010

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Revision as of 14:08, 13 October 2010

Hydrophobofilm
pushing coatings into a greener future

Week 25, Arend Jan

Control restriction of pNZ8901-bbs with biobrick site enzymes. All were also cut with XhoI to check the orientation of the sites (in case the sites in the original plasmid were not removed).

- 10ul plasmid
- 1.5ul buffer G (XbaI, SpeI) or R (PstI)
- 0.5ul biobrick enzyme (XbaI, SpeI, or PstI)
- 0.5ul XhoI
- 2.5ul MQ

Clones 1 and 2. + = cut with XhoI, P+ = cut with PstI and XhoI, S+ = cut with SpeI and XhoI, X+ = cut with XbaI and XhoI

alt text

Because of the added XhoI a ~270bp fragment should be present. This is the case in all restrictions. All PstI restrictions give an unexpected 3 bands. The plasmid is ~800bp larger than the clone manager file but this was not considered a problem. It is likely that there is a PstI site in the unknown sequence. This site will have to be deleted before we can use this plasmid in multi-biobrick cloning steps.

As an extra control a restriction was done with BamHI which was used to linearize the plasmid after inserting the biobrick sites. Together with XhoI this should give a ~300bp band.

- 5ul plasmid
- 1.5ul buffer G
- 0.5ul BamHI
- 0.5ul XhoI
- 7.5ul MQ

Gel 23-06-10

@@ Line 13: / Line 13: @@
 - 2.5ul MQ </pre>
-[[Image:21-06-10-1gn.jpg|200px|thumb|left|alt text]] [[Image:21-06-10-2gn.jpg|200px|thumb|left|alt text]]
+[[Image:21-06-10-1gn.jpg|200px|thumb|left|Clones 1 and 2. + = cut with XhoI, P+ = cut with PstI and XhoI, S+ = cut with SpeI and XhoI, X+ = cut with XbaI and XhoI]] [[Image:21-06-10-2gn.jpg|200px|thumb|left|alt text]]
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