This lab was pretty neat! We went through a step-by-step process to figure out a different transformed version of bacteria. For instance we found out if they were more similar to the original non-transformed E. coli colonies. The genetically transformed bacterial cells were transformed after we did these steps (some pictures shown above):
- Label one closed micro test tube +pGLO and another -pGLO
- Open tubes and use transfer pipet to transfer 250 ul of CaC12
- Place tubes on ice
- Pick up colony of bacteria with sterile loop
- Spin loop in +pGLO tube until colony is dispersed
- Place back in ice
- Repeat with -pGLO tube
- Examine pGLO plasmid DNA solution with UV lamp
- Immerse new sterile loop into the plasmid DNA stock tube and withdraw a loopful
- Mix loopful into the cell suspension of +pGLO tube
- Close and return to ice
- Close -pGLO tube but DO NOT add plasmid DNA
- Incubate tubes on ice for 10 minutes!
- Put both tubes in water bath at 42 degrees C for 50 seconds
- Place back in ice for 2 minutes
- Remove and open a tube and use new sterile pipet to add 250 ul of LB nutrient broth
- Reclose it and repeat with other tube
- Incubate tubes for 10 minutes
- Pipet 100 ul of the transformation and control suspension onto the plates
- Spread suspensions evenly around surface
- Stack plates and tape together
- Stack and place upside down in 37 degrees C incubator until next day
When we were all done the results were pretty cool! It came out glowing all purple and what not! :) I was surprised because I didn't really think it would look all that cool, but I guess all those steps paid off! It looked awesome! :)
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