The expression of proteins in cellular models or bacteria is quite common in order to study their function and structure or for large scale production.

To make a bacteria express a certain protein, the cells must take up the DNA sequence that encodes for that protein. Because proteins can be modified after production by some cell-specific mechanisms, the final protein produced by a bacteria is not necessarily exactly the same as we would find in the original environment.

Not all bacteria can take up DNA, the bacteria who can do this are called competent bacteria, and to the process of inserting foreign DNA into a bacteria, we call transformation.

About a year ago, I had to do transformation of some competent bacteria. I did not want to express the protein, I needed to be able to distinguish a sequence of DNA that had been mutated from its normal version. We were hoping that in every four bacteria: one would have taken up the mutated version, one would have the control version and the other two would have  a mix of these two sequences. Because bacteria grow very fast, overnight I would have more than 100 colonies, from which I could select 10-20 and hoping “Mendel was on my side” send this for sequencing and voilà: I would know the exact mutation I had introduced in that DNA.

I have discovered the hard-way that even competent bacteria are not so competent if the scientist is not bright. After two months of failing to see bacterial colonies, asking for help from everyone I could find in the lab, making them read my protocol, comparing it to their protocol to no avail, the issue started bugging the professor. On a hallway conversation he says “Are you sure the water bath is at 42 C?” To which I reply “which water bath? I am using the thermo-block.”. It so seems that there is a preference for water baths over heating-blocks among the older generation of scientists. Is this well supported? The short answer is: of course not!

But there are differences between water-baths and heating blocks

Water baths need a very big amount of water to function, thus once they reach a certain temperature they will be very stable in maintaining it constant throughout the entire surface. This means that every single sample will be exposed to the same temperature as the one next to it and that every single particle of a given sample will be at that temperature.

Heating blocks on the other hand are constantly giving away heat, the temperature along its surface varies a bit (I never measured but I wouldn’t expect it to be much more than a few tenths of degree), also the temperature from the bottom of the tube to the upper part should vary a bit.

Is this sufficient to make a difference in getting colonies of bacteria growing?

In our lab, we will never know: people that used to use water bath still use them, people who used heating blocks still use them. And I, I have converted! Working experiments are better than non-working ones!

In any case, how does putting bacteria at 42 C help them taking up DNA?

Basically, we “heat-shock” them: the bacteria are on ice for 30 minutes, all cold and stiff. Then they go directly into 42 C, which makes them more comfortable. Suddenly, after 50 seconds, they go back to ice. As bacteria membranes are made of lipids, and lipids are very fluid, the changes in temperature allow for pores to be open in the cell membrane, through which DNA can move in without resistant.

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