褐飞虱中肠内膜结合短肽P2S的筛选

Screening of a peptide P2S binding to the midgut inner membrane in the rice brown planthopper, Nilaparvata lugens (Hemiptera: Delphacidae)

【目的】筛选与褐飞虱Nilaparvata lugens中肠内膜相结合的短肽,探索能阻断褐飞虱传播水稻病毒的新方法。【方法】通过改良的膜饲喂法和噬菌体短肽文库展示技术,模拟褐飞虱正常取食的行为,筛选能够与其中肠内膜结合的短肽。将筛选到的短肽与加强绿色荧光蛋白进行融合表达,经纯化后,将融合蛋白饲喂给褐飞虱若虫,通过荧光显微镜观察饲喂后若虫的中肠。【结果】使用改良的膜饲喂法,16h后褐飞虱若虫的取食率达到91%,24h后达到95%。经形态学验证,确定了一种短肽与褐飞虱中肠内膜具有结合活性,并将其命名为P2S。【结论】改良了针对褐飞虱的膜饲喂法,筛选并验证了能够与褐飞虱中肠内膜结合的活性短肽P2S。研究结果为阻断相关的水稻病毒与褐飞虱中肠内膜间的相互作用研究提供了科学依据,也为褐飞虱新型膜穿孔毒素蛋白的研究提供了理论基础。

[ Aim] To screen peptides which may bind to the midgut inner membrane in the rice brown planthopper, Nilaparvata lugens, so as to explore the new methods for blocking N. lugens to transmit rice viruses. [ Methods ] Trophic behavior of nymphs of the rice brown planthopper was imitated with improved membrane was fed to the fusion feeding method. Through the use of this feeding method, the phage peptide library solution planthopper nymphs to screen peptides which may bind to the midgut inner membrane. Then, expression of peptide and enhanced green fluorescent protein was conducted, and the purified fusion protein was fed to nymphs of the planthopper. To confirm the binding activity of screened peptide, the green fluorescence of fusion protein in the midgut of the nymphs was observed under the fluorescence microscope. [ Results] The feeding rates of nymphs of the planthopper detected by improved membrane feeding method at 16 h and 24 h after treatment reached 91% and 95%, respectively. By observing the green fluorescence from planthopper gut, one peptide, named P2S, was determined as the planthopper gut inner membrane binding peptide. [ Conclusion ] Membrane feeding method was well improved for the rice brown planthopper. A peptide P2S which may bind to the midgut inner membrane of the planthopper was effectively screened and its binding activity was confirmed. The results will provide a possibility of blocking interaction between rice viruses and the midgut inner membrane of the rice brown planthopper. Furthermore, the peptide P2S may be used for further study of the membrane insertion toxins binding to proper receptors in the midgut of the rice brown planthopper.