Week of 6/03/24-6/06/24 Electrophoresis and Plasmind (PRAD1) Extraction

Returning to lab on Monday, we found that another lab mate's competent cells we had utilized for our project was contaminated so we had to scrap all of our bacterial specimens that was used with those cells. This set us back quite a bit, however we were able to bounce back. In the future, we intend on using our own supply of any cells we may need in order to reduce the risk of contamination again due to technical errors. It was established that we will be making R2A media every Monday, preferably 250ml of such in order to comfortably have a steady line of media. On Tuesday, we took the OD value of Sonora using R2A as our blank, with which we received an OD value of 1.63. On Wednesday we inoculated 600 micro liters of our bacterial culture with colony PCR in order to verify that our transformation of Sonora was successful. E. Coli was our positive control, while Sonora would be our negative control. This is because E. Coli contains the plasmid PRAD1 already, which is what we are trying to transform Sonora with. On Thursday we loaded our gel into the electrophoresis process, garnering results of two faint red bands being observed within ladder 4 and ladder 6. This means that our verification of a successful transformation is inconclusive. An area for concern is the fact we had failed to even use the Gel Red dye during the PCR process, so we should not have seen anything at all regardless. This could mean there is contamination present within our samples. Moving forward, we will be enforcing more critical analysis of material use and ensuring more gram stains are performed more often. 

Desiccated (Zombie) Sonora under microscope.







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