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Question about Comparing E.coli Strains for Transformation Efficiency

Posted: Wed Nov 29, 2023 3:55 pm
by moonagedaydream
Hello!

I am doing a science experiment comparing the transformation efficiencies of various strains of E. coli. For my experiment, I am using the pGLO plasmid from BioRad as my plasmid being cloned, and the different strains of E. Coli I am comparing are E. Coli K-12, E. coli B, and E. Coli DH5a. My results show that E.Coli K-12 had the highest rate of transformation efficiency, E.Coli DH5a had the second best, and E. Coli B had the worst. I understand why Ecoli. B has the worst transformation efficiency; historically, it has never been the best at transformation. However, I am a little confused why E.Coli K-12 has such a higher rate of efficiency than E.Coli DH5a, because E.Coli DH5a has so many mutations that are supposed to make it the most efficient. I have tried searching online for reasons I may have gotten these results, but everything I found is very vague, describing how E. coli DH5a might simply be more sensitive to prep conditions, or that the strains just have different builds. I'm looking for some help on interpreting my answers! Any specific facts on why I got these results would be very helpful - like, I understand how my results are different because the strains are composed of different proteins, but I want to know what specifically! I'll provide more background on my project below that may or may not have influenced my results, and would be helpful.

The kit from BioRad, which the pGLO plasmid came with, suggested that K-12 be used as the bacteria. I presume that's because the pGLO is more suited for a K-12 strain, but I'm still wondering why!

The DH5a was the only bacteria that came already competent and frozen, the K-12 was lyophilized, and the B was growing on an agar plate. I made sure that I prepared them to contain the same volume of cells despite their different forms

I used the same heat-shock procedure for all strains of bacteria, keeping them on ice/in the hot bath and growing on agar plates for the same amounts of time.

I used agar plates with ampicillin to determine my yield quantity because the plasmid has ampicillin resistance, so presumably, the more colonies on a plate, the better the bacteria's transformation efficiency is. However, I also ran trials on agar plates that did not have ampicillin, and all the bacteria grew just fine, so I know it's not an issue with the Ecoli itself.

Thank you for all of the help. Anything would be appreciated!!