アセチルコリンエステラーゼ (Acetylcholinesterase) · 12日 6月 2026

Furthermore, I set out to examine whether the point mutations at three amino acid sites, which the graduate student and I had detected and which French researchers had previously reported as important for organophosphate resistance, actually contributed to resistance. The outline of the experiment was as follows. I established a mass population from individuals newly collected in 2006, allowing various genotypes to be generated through combinations of the multiple alleles identified. I then...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 11日 6月 2026

Contribution of point mutations to organophosphate resistance Of course, I had to consider the possibility that resistance genes other than acetylcholinesterase were present in the population (indeed, the Katsunuma population also possessed a resistance factor on the second chromosome). Furthermore, it was possible that point mutations at amino acid sites in acetylcholinesterase that had not yet been analyzed should not be overlooked. Therefore, even though I was exploring genetic variation for...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 10日 6月 2026

This may not seem particularly significant to those who are not interested in the topic, but since deciding to study insecticide resistance, I have conducted my research based on the hypothesis that the possibility of multiple resistance factors in insect populations should not be ruled out. In that sense, I feel I was able to obtain fairly satisfactory results. I was able to begin my experiments on acetylcholinesterase immediately after returning from Edinburgh, and within the limited time of...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 09日 6月 2026

Direct sequencing of partial PCR products of the acetylcholinesterase gene revealed that at least four types of alleles—including a susceptible allele—exist in the natural population of D. melanogaster in Katsunuma, depending on the presence or absence of amino acid substitution mutations at the three sites reported by French researchers to be particularly important. We had previously discovered that the Katsunuma population possessed resistance factors on both the second and third...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 08日 6月 2026

Confirmation by direct sequencing On the other hand, if an individual is homozygous for point mutations at each of these three sites, the genotype can be interpreted as representing a single allele, at least with respect to these sites. Therefore, I used a method known as direct sequencing to examine DNA sequences from individuals identified as homozygous at these sites by allele-specific PCR. This was done to confirm that these alleles had not been misidentified by allele-specific PCR and that...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 07日 6月 2026

Nevertheless, the results differed from what I had originally expected, and various difficulties arose with the method I had been using up to that point. Originally, the allele-specific PCR method had been developed under the assumption that there was a single type of resistant allele (namely, the one carried by the resistant lines analyzed by the graduate student) along with a susceptible-type allele. However, when multiple alleles exist, as in this case, it becomes difficult to determine the...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 06日 6月 2026

When I performed allele-specific PCR using isofemale lines established from Drosophila newly collected in 2006, the existence of other resistant alleles had never occurred to me. Therefore, when I saw the electrophoresis patterns of the PCR products, which were contrary to my expectations, I felt a strong sense of excitement—almost a feeling of “Wow, this is it!”—that I would very much like to convey, although I suspect that only someone who has actually performed such experiments can...

アセチルコリンエステラーゼ (Acetylcholinesterase) · 05日 6月 2026

Problems with allele-specific PCR The resistant strains #609 and #1465—which I established from natural populations of D. melanogaster collected in Katsunuma in 1997 and 1998, when I was a graduate student—both harbored resistance-associated point mutations at three amino acid sites considered particularly important. Therefore, at least with respect to these three sites in the acetylcholinesterase gene, only a single type of resistance allele was detected. At that time, there seemed to be...