Team:UofGuelph/Contributions
Registry Contributions
2018 Registry Submissions
Sequencing of BBa_K747047
Background
iGEM Guelph's 2018 contribution to the registry was the sequence analysis of a BioBrick located in the 2018 Distribution Kit.
BioBrick K747047 was located in Plate 2, Well 12O. Our team chose to sequence this part as the Distribution Kit listed it as a Bad Sequence and for our team’s first characterization of a Registry Part, we thought that this part was a good candidate. Sequencing preparation was performed by our team, while the sequencing was performed at the Laboratory Services at the University of Guelph.
Methods
A PCR was set up according to the following recipe:
| Component | Volume (µL) |
|---|---|
| H2O | 20.4 |
| Phusion Buffer | 8.0 |
| dNTPs | 0.8 |
| VF2 | 0.2 |
| VR | 0.2 |
| BBa_DNA | 10 |
| Phusion | 0.4 |
| Total | 40 |
PCR Cycling Parameters:
| Step | Temperature (°C) | Time (minutes) |
|---|---|---|
| Initial Denaturation | 98 | 2 |
| Denaturation | 98 | 1 |
| Annealing | 60 | 1 |
| Extension | 72 | 0.5 |
| Final Extension | 72 | 10 |
"Denaturation", "Annealing", and "Extension" steps were repeated 34 times.
10µL of the PCR reaction was then run on a 1% agarose gel at 130V for 10 minutes to confirm presence of an amplicon.
The samples were then sent for sequencing via Sanger Sequencing.
10µL of the PCR reaction was then run on a 1% agarose gel at 130V for 10 minutes to confirm presence of an amplicon.
Results
Raw Sequence Data (trace files) can be found through this link
The sequences were then uploaded to the registry as part of our submission for our team to earn the Bronze Medal.
