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| probability that the E. coli absorbed the plasmid. The tired but excited young | | probability that the E. coli absorbed the plasmid. The tired but excited young |
| researchers went home with fingers crossed.</p> | | researchers went home with fingers crossed.</p> |
| + | |
| + | <h3>Thursday (06/08/17)</h3> |
| + | <p>When they entered the lab on the second day the plates showed growth |
| + | with distinctions of individual |
| + | colonies. Mentor Mike Goodson |
| + | ecstatically let the students know about the |
| + | significance of the growth and students were |
| + | relieved that their plates had grown! These |
| + | colonies were used to complete the PCR |
| + | protocol. The master mixture, (protocol tab) and |
| + | LB broth in round bottom falcon tubes were left |
| + | in the incubator (37°C), overnight. The plates |
| + | were put in the refrigerator (4°C) to preserve the |
| + | bacteria for future use. This protocol was used |
| + | to confirm that E.coli JM109 had absorbed the |
| + | plasmid PRhl_GFPa1</p> |
| + | |
| + | <h3>Friday (06/09/17)</h3> |
| + | <p>For the last day of the week, the students’ goal was to confirm the E. Coli |
| + | had absorbed the plasmid and then perform a MiniPrep Kit to take the |
| + | plasmid out of the bacteria. To accomplish this, they first learned how to |
| + | create a gel and how to use the electrophoresis. The electrophoresis sends the DNA |
| + | down the gel using a positive and negative charge. Under a blacklight you can see if |
| + | the DNA is present. Next, the students use a QIA prep spin Miniprep Kit to take |
| + | plasmid out of bacteria. The students placed one nanodrop onto spectrophotometer |
| + | to measure how pure the DNA extracted is. For future experiments, the plasmid DNA |
| + | would then be sent to an offsite lab to be sequenced.</p> |
| + | |
| + | <h2>Week 3: June 12-16</h2> |
| + | <h3>Monday (06/12/17)</h3> |
| + | <p>The students went to a group meeting within their labs. Students |
| + | streaked bacteria onto agar plates and put back into the incubator. The |
| + | rest of the day was spent collaborating on website design and graphic |
| + | design. Two coding novices, Annie and Jason, worked on learning html, CSS, and |
| + | Javascript. Graphic designer Tina created a new mascot microbe. Assistant graphic |
| + | designer and artist Annie created a drawing of the original Wright Flyer and assisted |
| + | in Photoshop drawing. Stenographer Hayley typed up protocols and helped organize |
| + | team documents.</p> |
| + | <p>The whole team met in the morning to discuss the Wiki and each group’s project. |
| + | The RX team observed the preparation and running of PCR. The team made LB |
| + | broth and agar. Later the team observed the purification of the PCR results and the |
| + | analyzation of the results with Qubit.</p> |
| + | |
| + | <h3>Tuesday (06/13/17)</h3> |
| + | <p>-The students removed the agar plates with bacteria from the incubator. |
| + | Two tubes were filled with LB broth, one labeled as a blank and one |
| + | containing the bacteria. Using a sterile toothpick, a single colony was picked |
| + | from each plate, and placed in the culture tube. An unused sterile toothpick was |
| + | added to the blank tube, to verify that the process was sterile. These tubes were |
| + | then shaken in an incubator for 3 hours.</p> |
| + | <p>The students conducted an experiment using culture growth to see if the bacteria |
| + | will turn green with the addition of the quorum sensing molecule to the bacteria.</p> |
| + | <p>The next morning, the plasmid should glow green indicating it is present.</p> |
| + | |
| + | <h3>Wednesday (06/14/17)</h3> |
| + | <p>The students retrieved the test tubes placed in the incubator the day before |
| + | and analytically checked the optical density and the fluorescence of the |
| + | colonies. Immediately afterwards, a visual check was performed as well, |
| + | confirming that the Colony + 3OC12 glowed green. The ecstatic students celebrated |
| + | as anticipated results showed through the light.</p> |
| + | <p>In the afternoon, the students were back at work again preparing a mini-prep on the |
| + | PLCD-GFPa1 (the plate streaked on Monday). The remaining DNA containing the |
| + | GFP was then placed in the -20o C freezer for storage. Having much more |
| + | experience the second time doing the mini-prep, the trailblazing students had a |
| + | much better grasp and spent less time with more accuracy.</p> |
| + | |
| + | <h3>Thursday (06/15/17)</h3> |
| + | <p>-Time to go back to work bright and early. A little after the crack of dawn, |
| + | the engaged students immediately went to work preparing a digest. The |
| + | scholars made a 15 uL digest along with a Master Mix. Interactive steps |
| + | included preparing a mini-prep, mixing with enzyme BSr61-HF, enzyme KpuI-HF, buffer Cutsmart, and water. The remaining solution was then placed in incubation for |
| + | 2 hours. In addition, students were excited to read and learn about plasmid cloning |
| + | by restriction enzyme digest and restriction endonuclease. With a desire to learn |
| + | more, the students trekked back to the lab and met with another qualified scientist |
| + | to assist in the digest. Once the digest was |
| + | done, the students made a gel and |
| + | electrophoresis to analyze the DNA. The |
| + | electrophoresis sends the DNA down the |
| + | gel using a positive and negative charge |
| + | and allows the students to see the amount |
| + | of base pairs, and in this case also allows |
| + | the students to see the cut DNA and |
| + | replacement. However, the young scientists |
| + | were very disappointed to find out the the |
| + | digest did not go as expected and DNA did |
| + | not get replaced as expected. Disappointed |
| + | but not defeated, the students immediately |
| + | went back to work and retraced their steps, |
| + | beginning the process all over again. |
| + | Things do not always work out in the lab |
| + | and the students were eager to restart and achieve favorable results.</p> |
| + | |
| + | <h3>Friday (06/16/17)</h3> |
| + | <p>-Immediately the next day, the eager students were right back at work. The |
| + | previous day's failure had fired them up and they were eager to do |
| + | everything carefully and quickly. The day began with a student-led mini-prep |
| + | in which the students were able to follow the protocol and do everything by |
| + | themselves. Afterwards, the students and mentors went out for a cool ice cream |
| + | treat. All 12 days of hard work deserved a celebration and the students and mentors |
| + | bonded over frozen custard and ice cream.</p> |
| + | |
| | | |
| </div> | | </div> |