Identification of the gene responsible for host-plant selection in Drosophila sechellia

对果蝇种间杂交种和黑腹果蝇缺陷型突变体对己酸的选择性进行了测试。3株缺陷型菌株在本试验中表现出己酸偏好。然而，现有的细胞学数据并不支持这三个缺陷之间存在重叠区域。由于细胞学数据不够精确，我们在分子水平上检查了这些缺陷的断裂点。PCR标记显示重叠区的存在，大小为80kb。根据黑腹毛虫基因组计划的数据，这个区域包含22个预测基因。通过RT-PCR，我们证实了所有这些基因都在杜氏藻中表达，这表明杜氏藻中负责任的基因改变影响的是基因产物的结构，而不是它的表达模式。假设该区域的黑腹夜蛾与黑腹夜蛾同源，我们通过筛选黑腹夜蛾粘粒文库来克隆相应的基因组片段，得到了覆盖大部分靶区的两个独立克隆。通过鸟枪测序确定其全序列，并与黑腹果蝇基因组序列进行比较，以确定致病基因。

Inter-specific hybrids between Drosophila sechellia and deficiency mutants of D. melanogaster were tested for their preference to caproic acid. Three deficiency strains showed caproic acid preference in this test. Available cytological data, however, did not support the existence of overlapping region among these three deficiencies. Since cytological data are not precise enough, we examined the break points of these deficiencies at molecular level. PCR markers revealed the existence of the overlapping region, and it sizes 80kb. According to the genome project data on D. melanogaster, this region contains 22 predicted genes. Using RT-PCR, we confirmed that all of these genes are expressed in D. sechellia, suggesting that the responsible genetic alteration in D. sechellia affects on the structure of the gene product rather than its expression pattern. Assuming synteny between D. sechellia and D. melanogaster in this region, we screened D. sechellia cosmid library to clone the corresponding genomic fragment, and obtained two independent clones that cover most of the target region. Complete sequence was determined by shotgun sequencing, and was compared with D. melanogaster genomic sequence to identify the responsible gene.