Studying plant genes with a paintbrush and a baking tin

Image by J. Murray. Used with permission.

A paintbrush and a baking tin might seem unlikely equipment for scientists to use to study plant genes but both feature in a study recently published in the journal Plant Cell. Intrigued? Let me explain…

Imagine for a moment that you are a research scientist studying how legume plants (e.g. peas, beans) set up friendly relationships with soil bacteria called rhizobia. The rhizobia provide the plants with much needed nitrogen. In return, the plant provides the rhizobia with carbohydrates and a home within the plant roots in special organs called nodules.

To enter the nodules, the rhizobia first have to infect into the root, which is a complex process with multiple stages. We currently only know about a few of the plant genes involved, so to identify more genes, you would like to start by getting an overall picture or “profile” of which genes are switched on (expressed) as the rhizobia infect into the root.

Bacteria (stained in blue) infecting into a root hair cell as seen under a light microscope. Image by the author.

Most of the studies that have profiled how large numbers of genes are expressed in plants have used samples from large pieces of plant tissue, for example from whole roots or leaves. However, rhizobia do not infect plant cells evenly across the whole root and most of the cells in the deeper layers remain uninfected, so if whole root samples are used it may be difficult to spot the genes involved in infection. The rhizobia start by infecting individual cells on the surface of the root called root hair cells (see image). How could you separate out the root hair cells from the rest of the root?

Some of my friends at the John Innes Centre, UK have addressed this question. Breakspear, Liu and colleagues (2014) froze the roots of the legume Medicago truncatula in liquid nitrogen to preserve the genetic material, and then removed the hair-like part of the root hair cells by brushing them with a small paintbrush (1). However, in the first attempts, the fragments of root hairs stuck to the sides of the container holding the liquid nitrogen so they switched to using a Teflon-coated (non-stick) baking tin, which allowed the root hair fragments to be transferred into tubes so the genetic material contained could be extracted for gene expression profiling (using fancier equipment).

Breakspear, Liu and colleagues used this method to study gene expression in the root hairs of Medicago truncatula at 1,3 and 5 days after they added rhizobia to the plants (1). They found that many genes were more highly expressed in these plants than in the control plants that had no infection.

The ARF6a gene and several other genes that were switched on during infection can also be switched on when plants are treated with the plant hormone auxin, which is known to have a role in making root nodules, but it has not yet been shown to be involved in infection. Further experiments on ARF6a showed that it is strongly switched on in infected root hairs only and M. truncatula plants missing the ARF6a gene had lower levels of infection than normal plants. These findings suggest that auxin is involved in promoting infection.

Many of the other genes that are highlighted in this study are involved in responses to other plant hormones, or communication with the rhizobia. These genes could be interesting subjects for future studies. The gene expression data is itself a useful resource for other scientists and has been deposited in an online database, The Medicago Gene Expression Atlas, which contains lots of other data from the various tissues and organs of M. truncatula in different situations.

This is just one example of how everyday items can find a new use in science experiments. Clingfilm (food wrap) and foil are essentials in most biology labs. Toothpicks, cotton wool, fabric netting and sewing thread are just a few of the everyday items I’ve used in the lab. Since problem solving by individual scientists is behind many methods used in science, it is perhaps not surprising that everyday items are involved.

Reference:

1. Breakspear, A., Liu, C. Roy, S., Stacey, N., Rogers, C., Trick, M., Morieri, G., Mysore, K.S., Wen, J., Oldroyd, G.E.D., Downie, J.A. and Murray, J. (2014) The root hair “infectome” of Medicago truncatula uncovers changes in cell cycle genes and reveals a requirement for auxin signaling in rhizobial infection. Plant Cell.