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Team:Macquarie Australia/Protocols/ArrivalofGBlocks
From 2012.igem.org
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| + | We had experienced delays in receiving our gBlock fragments due to high GC content in the designed fragments which would have caused problems with hairpin loop formation and thus we had to reduce the GC content. By reducing the GC content of certain fragments, we were forced to change the 30 bp overlap sequence in order for these sequences to overlap during Gibson Assembly. At one stage, Deino_A contained 80% GC content and thus had to be altered in order for synthesis by IDT to begin. | ||
Revision as of 08:58, 31 August 2012
Ordered gBlock fragments are nearly here!
Soon our gBlock fragments will arrive from IDT. On Tuesday (4/09/2012) we will perform Gibson assembly will assemble the following four biobricks:
- +T7 promoter Agrobacterium tumefaciens bacteriophytochrome
- -T7 promoter Agrobacterium tumefaciens bacteriophytochrome
- +T7 promoter Heme oxygenase
- -T7 promoter Heme oxygenase
Click on each above to see the corresponding fragment nucleotide sequence and fragment properties.
| BioBrick name | Fragment(s) |
|---|---|
| T7 Heme oxygenase | Hemo_T7_A + Hemo_B + Plasmid |
| Heme oxygenase (T7 minus) | Hemo_A + Hemo_B + Plasmid |
| T7 Agrobacterium | Agro_T7_A + Agro_B + Agro_C + Agro_D + Agro_E + Plasmid |
| Agrobacterium (T7 minus) | Agro_A + Agro_B + Agro_C + Agro_D + Agro_E + Plasmid |
Once the Deinococcus Radiodurans fragments arrive we will assemble them using Gibson assembly. Due to the large sequence of the D. radiodurans bacteriophytochrome, the fragments will be assembled without a T7 promoter. The designed sequences of the 5 fragments can be viewed here
| BioBrick name | Fragment(s) |
|---|---|
| Deinococcus radiodurans bacteriophytochrome | Deino_A + Deino_B + Deino_C + Deino_D + Deino_E + Plasmid |
We had experienced delays in receiving our gBlock fragments due to high GC content in the designed fragments which would have caused problems with hairpin loop formation and thus we had to reduce the GC content. By reducing the GC content of certain fragments, we were forced to change the 30 bp overlap sequence in order for these sequences to overlap during Gibson Assembly. At one stage, Deino_A contained 80% GC content and thus had to be altered in order for synthesis by IDT to begin.
