IPTG Induction vs Auto-Induction

by Simon Currie

IPTG and auto-induction are two ways to induce protein expression in bacteria. They work similarly, but have different trade-offs in terms of convenience. While IPTG induction is usually the default technique, there are situations where auto-induction are necessary.

I had been optimizing my protein’s expression for months and I thought that I had already tried nearly everything that could be done. Still, I was getting poor yields and less than desirable purity at the end of my purifications. And that’s when I came across auto-induction.

Auto-induction is an alternative method for inducing protein expression in bacteria for plasmids with the lac operon. Auto-induction uses defined media, and a programmed shift from glucose to the lactose metabolism to trigger protein expression (Studier, 2005).

IPTG and auto-induction are two ways to induce protein expression in bacteria. They work similarly, but have different trade-offs in terms of convenience. While IPTG induction is usually the default technique, there are situations where auto-induction are necessary.

This article compares auto-induction and IPTG induction. While IPTG induction is usually tried first, auto-induction is a great choice if IPTG didn’t work very well, or if you are doing a lot of fermentations in parallel.

Article Table of Contents

What is auto-induction?

IPTG or auto-induction?

References

What is auto-induction?

Auto-induction refers to a method of inducing protein expression when bacteria shift from glucose to lactose metabolism.

Bacteria preferentially use glucose as their primary energy source and will consume all of the glucose in the media before shifting to lactose (Blaiseau and Holmes, 2021). Once glucose is expended and cells shift to using lactose as their energy source, then allolactose, a metabolized version of lactose, is generated, and that will trigger your protein’s expression by inactivating the lac repressor (Figure 1).

For that reason, auto-induction (and IPTG induction) only works with plasmids that have the lac operon.

Figure 1. Allolactose inactivates the lac repressor enabling transcription to proceed.

So, auto-induction timing is programmed by controlling the amount of glucose and lactose that goes into the media at the start.

Auto-induction requires more upfront work to make the media. But, once you start growing your culture it’s a lot less work since you don’t need to obsessively track the growth of your culture (OD₆₀₀) to time when to add IPTG, or optimize how much IPTG you’re adding.   

IPTG or auto-induction?

There are two scenarios where I suggest trying auto-induction:

1)    You’ve already optimized IPTG induction, and still the protein quantity and/or quality is not sufficient.

2)    You’re doing a lot of fermentations in parallel and it will be a lot of work to use IPTG to induce each one.

To go back to the story, when I finally figured out that auto-induction worked wonders for the key protein I had been trying so hard to get (Currie et al, 2017), I thought auto-induction was going to be my go-to protein expression technique for every protein that I would work with. Over time, however, I saw that for most proteins auto-induction and IPTG induction were pretty comparable. In fact, sometimes the protein expression was even a little bit lower with auto-induction.

Because of this experience, I usually try IPTG induction first, then move onto auto-induction for the few protein constructs that have poor expression with IPTG.

The other consideration in all of this is ease-of-use. Auto-induction requires making defined media in advance, including concentrated solutions of the glucose and lactose solution, salts, etc. Most of this work is done up front, and these stocks are used for multiple expression cultures. However, the concentrated salt solution is, in particular, a pain to work with. The concentrated salts crystallize at room temperature, so each time you have to heat it up to get the salts back into solution. This takes time and can make a bit of a mess on your hot plate or in your microwave.

By comparison, the media for IPTG induction is usually just Luria Broth (LB) or Terrific Broth (TB), and is quite easy to make and is very soluble.

The trade-off is that for auto-induction, there is no need to carefully track culture growth or optimize IPTG induction. Instead, you just start the growth, if desired turn down the temperature when you’re nearing expression (roughly around an OD₆₀₀ of 1.0), and let it grow, ~ 12 – 24  hours after that.

Given these differences, IPTG induction is easier when you’re doing a few cultures in parallel, whereas auto-induction is more convenient if you’re doing many cultures in parallel. That’s because each culture will grow at slightly different rates, and with auto-induction you won’t have to track that growth and add IPTG to all of the cultures at just the right time for each individual culture.

In fact, enabling expression of many cultures in parallel was a foundational motivation for the development of auto-induction in the first place (Studier, 2005).

To be clear, both IPTG induction and auto-induction are useful tools in your protein expression arsenal (Figure 2). Give both a try with a range of protein constructs to see which method you prefer.

Figure 2. Comparing auto-induction and IPTG induction.

Regardless of whether you’re using IPTG induction or auto-induction, we have reliable BL21 competent cells to support your protein expression needs. See below for lots of additional great GoldBio products and supporting resources that will have you purifying protein in no time!

References

Blaiseau, P. L., & Holmes, A. M. (2021). Diauxic Inhibition: Jacques Monod's Ignored Work. Journal of the history of biology, 54(2), 175–196. https://doi.org/10.1007/s10739-021-09639-4

Currie, S. L., Lau, D. K. W., Doane, J. J., Whitby, F. G., Okon, M., McIntosh, L. P., & Graves, B. J. (2017). Structured and disordered regions cooperatively mediate DNA-binding autoinhibition of ETS factors ETV1, ETV4 and ETV5. Nucleic acids research, 45(5), 2223–2241. https://doi.org/10.1093/nar/gkx068

Studier F. W. (2005). Protein production by auto-induction in high density shaking cultures. Protein expression and purification, 41(1), 207–234. https://doi.org/10.1016/j.pep.2005.01.016

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auto-induction IPTG protein expression Protein Induction

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