Biotechnology student IJse Boogaard silences plant genes to investigate their effect on thrips.
‘Plants obviously cannot move, so they have developed other defence mechanisms against threats such as herbivory – for example by secreting compounds. Herbivorous insects dislike these compounds, so in turn they have developed molecules to suppress plant defences’, says IJse Boogaard. The recently graduated HBO biotechnology student examined, for his final-year research project, the arms race between thrips (Frankliniella occidentalis) and tomato and pepper plants (Solanum lycopersicum and Capsicum annuum). ‘Thrips are only 2 to 3 millimetres long, so the small amount of feeding damage they cause is something most plants can tolerate. Even so, they are very dangerous to plants, because thrips can transmit viruses. In particular, tomato spotted wilt virus causes a great deal of problems’, Boogaard explains. In short, thrips feed on infected plants and then spread the virus to other plants via their saliva.
Interaction
Boogaard focused on one aspect of the battle between plants and insects: the interaction between plant proteins and the anti-defence molecules produced by insects. ‘Whether thrips produce such molecules is not entirely certain, but it is expected that they do. In the research group at the University of Amsterdam where I did my placement, a number of candidate molecules had already been identified, along with a number of candidate plant proteins with which they might interact. I temporarily silenced four genes encoding such plant proteins to see what effect that had on the plants and on thrips’, Boogaard explains.
Harmless
To do this, Boogaard used the so-called virus-induced gene silencing technique. ‘You introduce a harmless virus into the plant, and into that virus you insert DNA from the gene you want to silence. The plant combats the virus by destroying all the RNA transcribed from the viral DNA, and now does the same for RNA corresponding to the piece of DNA you have added. The effect is temporary: it takes effect four weeks after treatment. You then have to carry out your experiments quickly, because two weeks later the effect has already worn off.’
For Boogaard, this meant mapping feeding damage and counting thrips eggs. The results are mixed. One silenced gene produces malformed tomato plants. Boogaard repeats his experiment with the model plant Nicotiana benthamiana, but those tobacco plants die altogether. Other genes alternately lead to more or fewer thrips eggs and, in one case, to less feeding damage.
‘It is not the case that tomato growers can immediately do something with these results. But it is useful for them to know whether a gene has positive or negative effects on the plant or on thrips. That knowledge can be used in breeding, for example’, Boogaard explains.
Snag
There is also a snag, because one crucial experiment fails for Boogaard: ‘Strictly speaking, all my data are worthless, because I cannot truly prove that I have silenced the genes. I wanted to use qPCR to visualise gene expression, but that kept failing. It went wrong during the isolation of RNA from plant material, though I still do not know exactly why. Since the control experiment – in which plants with a silenced gene turned white – did work, we expect that the silencing was successful elsewhere too, but unfortunately hard data are lacking.’
Title: Research report: the impact of host plant proteins on the feeding and reproductive behavior of Frankliniella occidentalis
Author: IJse Boogaard
Category: bachelor’s thesis
Programme: bachelor’s in biotechnology, Inholland University of Applied Sciences; Assessors: Nina Guarneri, Maaike Boersma
Grade: 7.5
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