![]() They ultimately succeeded in identifying the time when the sex difference of this horn appeared. Upon this discovery, the researchers then looked for a boundary between the time when the horn can be grown by the suppression of transformer gene and the time when the horn cannot be grown, by suppressing this gene in female larvae using RNAi taken from various stages during larval-pupal development. They found that when this gene is suppressed during the larval stage, male-like horns are formed in the female beetle. The research team subsequently revealed that the sex of the beetle is controlled by a gene called Transformer. Next, referring to the mechanism of sex determination in Drosophila, they examined how the sex of the beetle was determined. Late larval stage (left), prepupal stage (center) and pupal stage (right). As a result of this, it became clear that it took about 36 hours after the reference point for the morphological differences between males and females in the horn primordia to appear. Using this as a reference point, the research group was able to create a stage table of horn formation along a time axis. "We found that head-rocking behavior was a sign of the beginning of the prepupal stage," Dr. ![]() Morita then video recorded a few weeks of the process of beetle larvae becoming pupae, and looked for signs that the prepupal stage had started. Morita developed a rearing method encompassing the beetle's late larval stage up until adulthood that used plastic tubes, but didn't require any soil. Furthermore, because the larvae and pupae live in the soil, it was particularly difficult to observe their development." Therefore, Dr. Shinichi Morita, said, "Unlike the model animal Drosophila, we did not have any basic information concerning these beetles, such as their developmental stages.
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