Blog #1 1/22/24 - 1/26/24

 Introduction

This week was the biotech boot camp where we had a basic introduction to lab safety. We learned about the protocol for entering, leaving, and being in the lab. When you enter the lab you leave any drinks in the red bin (we are not allowed to drink anything), then you wash your hands and sign into the lab book. You also need to clean your table with fantastic. While being in the lab you should keep your space clean, be mindful of others, wear your goggles, and write in your lab notebook. When you’re leaving the lab you should clean and put everything away in your area. Write your final note in your lab notebook about what you did and plan for next time. Then wash your hands and sign out of the lab book. We learned some general rules like giving Chad and Jonathan 2 or 3 days in advance when you will need something. Along with always asking questions when you are unsure, never opening anything without consulting someone about it, and doing all your research outside of the lab. We went through different tools in the lab for example scales, flaskes, incubators, etc. We also practiced using pipettes. I also assist Alex with some gram-staining, plating, and other things.  


Methods

On Thursday we were testing the survivability of E coli. under UV radiation stress. We started by using the nanodrop to test the concentration of bacteria in different flasks. The process starts with cleaning the sensors on the nanodrop machine. After we proceeded to zero (or blank) it, To do this we put 2 microliters of LB media that the bacteria grew in using a 2-20µl pipette. After blanking the nanodrop we clean the sensors again (we have to clean the nanodrop every time we test something along with changing the pipette tip to avoid contamination) then we put a 2 µl drop of flask A on the pedestal and write down the result. Something went wrong with the first test of flask A. After the first test, we clean the nanodrop sensor and pedestal, then add 2 µl of flask B. We did the same process with sample C and then tested flask A again and had normal results.


After we did a gram-staining for all three flasks A, B, and C. we started by labeling three glass slides with A, B, and C and drawing a circle to indicate where the bacteria will go. After we turned on the bunsen burner, then we sterilized the loop by putting it in the flame. After you put the loop in the broth you put the loop on the glass slide where the circle is and spread the broth. After doing all three slides you let them air dry and then heat seal them by swiping each slide over the flame five times. After, to do the stain you put drops of crystal violet on each slide, and wait one minute then wash it off with deionized water. Then we add iodine to each slide wait one minute then wash it off with deionized water. After washing each slide with alcohol then quickly wash it off with deionized water. The final step is adding safranin for one minute, washing it off with deionized water, and then drying the slides with blotting paper. After the staining, we were able to look at the bacteria under the microscope. 


After viewing the three samples under the microscope we decided to use the broth in flask B in the experiment where we will be testing the survivability of E. coli against UV radiation rays. To expose the bacteria to UV radiation we needed to use the ultraviolet crosslinker. To start we plan out the experiment and measurements, we grab 7 little tubes and label them 1-7. We fill each tube with 900 µl of LB broth each tube. Then we Zapp 100  µl of the broth at 70 mJ/cm2 for 1 minute and then add it to tube one. Then we do the same but Zapp for two minutes and then add to tube 2. After we then we Zapp 100  µl of the broth at 60 mJ/cm2 for 1 minute and then add it to tube 3. Then we do the same but Zapp for two minutes and then add to tube 4. Then we Zapp 100  µl of the broth at 50 mJ/cm2 for 1 minute and then add it to tube 5. Then we do the same but Zapp for two minutes and then add to tube 6. For the final tube, we will zapp 100 µl of broth for two minutes at 90 mJ/cm2  as a control because  90 mJ/cm2 is the value where E coli. Begins to die. After we plated each of the samples onto agar plates and put them into the incubator. 

 

Results

The results of the nanodrop test of flask A were unusual. The graph that showed up was inverted where it went up instead of down. The concentration was -0.43. The other flask results were usual but we decided to redo the nanodrop test of flask A. The result of the second reading of flask A was 2.78. 

Figure 1.


The results of the gram stain were unusual. For flask A we did not see a lot of the bacteria but it was a gram-negative result. The results for flasks B and c were better and we had a clearer picture of the bacteria, which was a gram-negative bacillus. We ultimately decided to use flask b for our experiment where we would zap  E coli bacteria with UV radiation rays. Flask B was zapped with different values of UV radiation for two different time intervals, then plated and put into an incubator. 


Conclusions

This was my first week in the lab on Monday and Tuesday we had a boot camp where we just learned the basic safety and general rules of being lab. I’ve had some experience already with being in a lab setting but I did like having a refresher on the safety protocol. It was a good boot camp I learned the rules of this lab which is important. Everyone was nice and willing to answer any question we had. They put in the effort to create a welcoming and secure environment, which is very appreciated thank you. On Thursday I had more hands-on experience about what goes on in the lab. Alex was very patient and explained any questions I had. I was a little nervous so was hesitant to do some of the activity. I am hoping that after a while of being in the lab, I will become more comfortable and confident doing experiments.


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