Team:UPRM/Experiments
Objectives:
- Perform digestion procedure according to the preset cloning plan.
- Fragments and vectors digestion in order to obtain the specific DNA to the ligation procedure.
- Prototype assembly
- Successful transformation of the prototype in Escherichia coli.
We have built a complete Cloning protocol in order to achieve our prototype assembly.
Wet Lab
First Stage
The main objective is to successfully digest the seven fragments necessary to complete the prototype assembly.
| Fragment | Stock Concentration |
|---|---|
| 1a | 200 ng/µL |
| 1b | 200 ng/µL |
| 1c | 200 ng/µL |
| 2 | 200 ng/µL |
| 3 | 200 ng/µL |
| 4 | 200 ng/µL |
| 5 | 200 ng/µL |
The fragments was digested using EcoRI and PstI restriction enzymes. Digestion reactions were calculated for each fragment. After digestion enzymes were inactivated at 20 °C (EcoRI) and 80°C (PstI). Then, an electrophoresis gel was prepared to analysed the digested fragments.
Protocol specifications:
Digestion Reaction
- Mix in a 1.5 mL microcetrifuge tube
- 5μl DNA (土200 ng/ μl)
- 0.5 μl PstI
- 0.5 μl EcoRI
- 1 μl enzyme buffer
- 3 μl ddH2O
- Incubate at 37 °C for 15 min to 2 hrs.
- Inactivate enzyme at 60 °C for 20 min and 80 °C for 20 mins.
10 μl Total Volume
Our ideal working concentration is over 100 ng/uL.
Figure 2: Fragments digestion. In the electrophoresis gel (agarose 1%) strong fragment concentrations are notable. To see a fragment in a gel a minimum of a 100 ng is needed. The observed results indicate that a concentration of 土150 ng were conserved along the process due to the bands brightness.
SynBio 101 Results
Surveys were used as the instrument to gather quantitative information that represents the general public’s knowledge about Synthetic Biology, Genetically Modified Organisms (GMOs) and agriculture. This helped us have a better understanding of how obstacles in these topics could be targeted using our prototype. After interviewing 53 people, in summary, their answers regarding their knowledge about Synthetic Biology, their consumption of GMOs, and if they think the field is being addressed adequately by the general public:
During the Operation 456 Workshops we wanted to track the improvement in knowledge of the 35 participants. For this, students were given pre and post tests. We compared the results of these tests and saw an increase of correct answers from 41.43% to 78.93%, showing an improvement of 37.5%.
Thanks to our sponsors
