Plant–herbivorous insect networks: who is eating what revealed by long barcodes using high-throughput sequencing and Trinity assembly
Plant–herbivorous insect networks: who is eating what revealed by long barcodes using high-throughput sequencing and Trinity assembly
复制标题
植物-草食性昆虫网络：使用高通量测序和 Trinity 组装，通过长条形码揭示谁在吃什么
DOI：
10.1111/1744-7917.12749
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发表时间：
2021
期刊：
影响因子：
4
通讯作者：
A. B. Zhang
中科院分区：
文献类型：
--
作者：
Zhang X.M.;Z. Y. Shi;S. Q. Zhang;P. Zhang;J. Wilson;C. K. Shih;J. Li;X. D. Li;G. Y. Yu;A. B. Zhang
植物与昆虫之间的相互作用是特定自然群落中所有生物最重要的生命功能之一。在食物网研究中，通常需要大量的样品来确定昆虫的饮食。以前，使用短DNA条形码的桑格测序和下一代测序（NGS），导致物种水平鉴定低;同时，桑格测序的成本对于元条形码和更多样品来说是昂贵的。在这里，我们提出了一种快速有效的测序策略，以识别鳞翅目幼虫和他们的饮食在同一时间，而不增加成本的Illumina平台上在一个单一的HiSeq运行，与长的多重元条形码（COI的昆虫，rbcL，matK，ITS和trnL的植物）通过Trinity组装（SHMMT）获得。同时，采用桑格测序（单个个体）和NGS（多食性）验证了SHMMT方法的可靠性。此外，我们表明，SHMMT方法是快速和可靠的，与最高质量的序列的5个DNA条形码的63个幼虫个体（54个物种）回收（全长的100%的COI基因和98.3%的植物DNA条形码）使用Trinity组装（放大到1015 bp）。幼虫饲料鉴定，95%是可靠的，其余5%失败，因为他们的肠道是空的。通过SHMMT方法鉴定的食物与我们收集期间幼虫取食的寄主植物100%一致。我们的研究表明，SHMMT方法是可靠的和成本效益的昆虫-植物网络研究。这将有助于昆虫寄主植物的研究，通常包含大量的样品。
Interactions between plants and insects are among the most important life functions for all organism at a particular natural community. Usually a large number of samples are required to identify insect diets in food web studies. Previously, Sanger sequencing and next generation sequencing (NGS) with short DNA barcodes were used, resulting in low species-level identification; meanwhile the costs of Sanger sequencing are expensive for metabarcoding together with more samples. Here, we present a fast and effective sequencing strategy to identify larvae of Lepidoptera and their diets at the same time without increasing the cost on Illumina platform in a single HiSeq run, with long-multiplexmetabarcoding (COI for insects, rbcL, matK, ITS and trnL for plants) obtained by Trinity assembly (SHMMT). Meanwhile, Sanger sequencing (for single individuals) and NGS (for polyphagous) were used to verify the reliability of the SHMMT approach. Furthermore, we show that SHMMT approach is fast and reliable, with most high-quality sequences of five DNA barcodes of 63 larvae individuals (54 species) recovered (full length of 100% of the COI gene and 98.3% of plant DNA barcodes) using Trinity assembly (up-sized to 1015 bp). For larvae diets identification, 95% are reliable; the other 5% failed because their guts were empty. The diets identified by SHMMT approach are 100% consistent with the host plants that the larvae were feeding on during our collection. Our study demonstrates that SHMMT approach is reliable and cost-effective for insect-plants network studies. This will facilitate insect-host plant studies that generally contain a huge number of samples.