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Am I Damaging My E. coli by Spinning at High Speeds?

Dear Aunt Yersinia,

A very annoying postdoc in our group keeps telling me off for spinning E.coli at 13K in a tabletop centrifuge. The postdoc claims that high speed damages cytoskeleton and this will reduce my transformation frequency. But I don’t believe her as the cells are cushioned by water during centrifugation. Can you tell me who is right?

                                                  Irene (PhD student)

Dear Irene,

I understand your sentiment – nobody wants to linger near a centrifuge for 10 minutes if she can spin cells in 1 minute. You are right – a bit. Nobody talks about the damage due to the movement through the liquid itself.

But if you jump out of a plane without a parachute, it’s not the fall that’ll kill you, but the meeting with planet Earth. A similar effect happens with centrifugation, and the higher the speed the harder the fall. Eventually the bacteria will meet the bottom of the tube and each other at the bottom with a considerable impact. Compacting under the centrifugal force will break the outer structures, for example flagellae and will damage the internal structures such as cytoskeleton and even DNA.

Gram positive bacteria are less sensitive to this because of thicker cell walls. However, gram-negative bacteria, including E. coli, will suffer. So if somebody studies bacterial cytoskeleton, motility or biofilm formation, they should forget about spinning at a maximum speed.

Despite, or maybe because of, being annoying – meticulous people are often grating – the postdoc is right. E. coli is susceptible to “over centrifuging”. Competent bacteria with their membranes treated in various ways to enhance their DNA uptake, even more so.

So what should you do? 3,000 RPM for 5 minutes mostly does the job. You do not need to exceed 4,000 RPM on a table-top centrifuge or any other to spin E.coli.  Yeast are much larger and 3 minutes at 2,500 RPM will be enough to sediment > 95% of cells, although the  exact result depends on your centrifuge (remember RPM does not equal RCF), so do try this at your own lab.

As for the boredom of waiting for the cells to spin, I recommend using this time to tidy your bench or a communal sink and you won’t notice how 10 minutes pass.

Happy centrifuging.

Useful Reading:

Peterson B.W. et al.(2012) Bacterial Cell Surface Damage Due to Centrifugal Compaction  Appl. Environ. Microbiol. 78:120–5.

7 Comments

  1. NB on March 20, 2019 at 3:49 pm

    Almost exactly the information I was looking for, but why did you put an RPM speed instead of RCF? You even reference they’re not the same thing, and the RPM in this article is not a useful number, so why not just tell us the actual RCF we should be using?

  2. Aparna on December 22, 2017 at 7:04 am

    I agree with Aneesh! 🙂

  3. Ana Tronco on November 29, 2017 at 1:14 am

    What speed is okay to use if I want to concentrate transformed
    E.coli to grow without affecting them too much ?

  4. Shankar Gurung on April 27, 2017 at 5:26 pm

    Really helpful answer for my research problem I was facing.

  5. Clara on February 7, 2017 at 8:58 am

    Interesting article! Any chance you could give the equivalent RCF’s for these RPM numbers? Or at least the kind of centrifuge you have in mind? Thanks in advance!

    • Adrienne Huntress on May 24, 2017 at 2:20 am

      I came to the comments to ask the exact same question! I am super curious to find out. =)

  6. Aneesh on November 15, 2016 at 11:19 am

    Whenever I have a weird practical question that the internet or written manuals cannot answer, bitesizebio is always the way to go! Thanks a lot to all the writers and editors!

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