Difference between revisions of "Template:Team:Groningen/CONTENT/LOGBOOK/tasA Digestion t20"
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Latest revision as of 02:05, 18 September 2015
Digestion BBa_K823023 and BBa_K823024 (t20)
In this experiment the BBa_K823023 and BBa_K823024 backbones were digested to afterwards ligate it with tasA.
Our goal is to overexpress TasA by adding a second copy of the tasA gene under the control of a salt inducible promoter. This overexpression will lead to a stronger biofilm when the biofilm is exposed to salt water.
The digestion of the BBa_K823023 and BBa_K823024 backbones was successful.
<a class="postscriptum protocol" href="https://2015.igem.org/Team:Groningen/Protocols_and_Protocols/restriction">Restriction</a>
00:00, 2 June 2015 - 00:00, 2 June 2015
The backbones BBa_K823023 and BBa_K823024 were digested by the following steps.
#
Component
Amount
1
Mastermix
28 µL
DNA (BBa_K823023)
2 µL
2
Mastermix
28 µL
DNA (BBa_K823024)
2 µL
Digestion mixtures.
The samples were digested for 2 hours at 37 °C. Afterwards they were loaded on 1% agarose gel with SERVA DNA stain G.
#
Component
Amount
1
DNA (tasA 4.2)
20 µL
6x buffer
4 µL
2
DNA (BBa_K823023)
25 µL
6x buffer
5 µL
3
DNA (BBa_K823024)
25 µL
6x buffer
5 µL
Samples for on the gel.
The gel was run on 100 V for ∼1 hour and the following bands were visible:
tasA at ∼800-900 bp
K823023 (∼5000 bp) including a band of its RFP insert (∼1100 bp)
K823024 (∼6000 bp) including a band of its RFP insert (∼1100 bp)
These band sizes match the in silico sizes.