Yes! I have allowed myself to get distracted. But don’t worry; the rabbit hole in question is scientific. The dye, fast scarlet, does curious things to my roots. A curiosity I have already written about is the dark zone; for some reason, the stain is excluded from a relevant part of the root. I have been trying various things to let the stain in. One of those was to introduce the dye into the agar medium in which the roots grow. That way, the roots and the dye would have a nice long time to cozy up together. As for penetrating the zone of blackness, the dye in the agar failed. But it did tip me headlong among the rabbits.
In that previous post, I recounted how a concentration of 0.001% fast scarlet had no effect on the growth rate of the root but stopped root hairs in their tracks. I said then that “someone should study this.” Apparently, that someone is me.
Figure 1 Fast scarlet affects root growth and root hair emergence. A: Growth rate of roots vs time for two control plates (open black circles and diamonds) and two plates where the growth medium was supplemented with 0.0003% fast scarlet (filled red circles and diamonds). Each plate had about a dozen seedlings and the roots grew inside the agar. B: Brightfield micrograph of a control root. Lots of root hairs. C: Brightfield micrgograph (same conditions as for B) of a fast scarlet-treated root. Arrows point to inicipient root hairs (“bulges). The root has a faint red tinge from the stain.
I set up a trial with a lower concentration: 0.0003% fast scarlet. Again, I made a pair of plates at that concentration and a pair of controls. I plated the seeds so that the roots would grow inside the agar; I measured growth rate from day 5 through day 9. In this experiment, the roots growing in fast-scarlet grew a little bit faster than controls (Fig. 1A). Yes, the effect is small, each of the symbol shapes represents the average for one plate and the two plates of each type (control: black symbols; fast scarlet: red symbols) grew a little differently. But in the paper by Charlie Anderson and crew who characterized fast scarlet as a useful dye for staining cellulose (read their paper here), roots growing on 0.001% fast scarlet grew a faster than wild type, again by a small amount. My result (albeit at a lower concentration) becomes a little more real by being consistent with theirs.
What about root hairs? See for yourself (Fig. 1B, C). If you look closely at Fig. 1C, you will see the slimmest of bulges where a root hair began to emerge and stopped. To help, I labeled two such bulges with arrows. The dye slammed root hairs ubiquitously: I looked at centimeters and centimeters of root hair length and saw no bulge that was any bigger, let alone an emerged hair. Weirdly, the bulges were larger on the higher concentration (0.001%) where—stained red—they looked like hemispherical welts. I am definitely going to repeat the trial at 0.001% because of course, repetition. But I’d like pictures!
My interest here is not entirely random. Root hairs stopped in their tracks at the bulge stage is a phenotype of a mutant called buzz (publication). The bald roots covered with stubs reminded Karen Sanguinet, who found it, of military haircuts. When she found this mutant, Karen was working in my lab; in her own lab now, buzz is very much in play. Now, another buzz phenotype is that the primary root grows faster than does wild type (even three-times faster in one background). To be sure, buzz is a mutant of the aspiring model grass species, Brachypodium distachyon (“brachy” among friends). The affected gene (probably knocked out by the mutation) is a cyclin-dependent cell division kinase; but the gene is a family member that has been flung far enough by evolution for its activity to be anyone’s guess.
Even if its growth rate promotion is modest, fast scarlet phenocopies two features of buzz. I am fascinated, enough to chase it down the rabbit hole—for a few meters. This past week, I set up a trial on an even lower concentration (0.0001%). Let’s see what happens…