Department of Anatomy & Neurobiology, School of Medicine, Medical Sciences Campus, University of Puerto Rico, San Juan, Puerto Rico.
Institute of Neurobiology, Medical Sciences Campus, University of Puerto Rico, San Juan, Puerto Rico.
PLoS One. 2021 Apr 9;16(4):e0249801. doi: 10.1371/journal.pone.0249801. eCollection 2021.
Crustaceans are major constituents of aquatic ecosystems and, as such, changes in their behavior and the structure and function of their bodies can serve as indicators of alterations in their immediate environment, such as those associated with climate change and anthropogenic contamination. We have used bioinformatics and a de novo transcriptome assembly approach to identify potential targets for developing specific antibodies to serve as nervous system function markers for freshwater prawns of the Macrobrachium spp. Total RNA was extracted from brain ganglia of Macrobrachium carcinus freshwater prawns and Illumina Next Generation Sequencing was performed using an Eel Pond mRNA Seq Protocol to construct a de novo transcriptome. Sequencing yielded 97,202,662 sequences: 47,630,546 paired and 1,941,570 singletons. Assembly with Trinity resulted in 197,898 assembled contigs from which 30,576 were annotated: 9,600 by orthology, 17,197 by homology, and 3,779 by transcript families. We looked for glutamate receptors contigs, due to their main role in crustacean excitatory neurotransmission, and found 138 contigs related to ionotropic receptors, 32 related to metabotropic receptors, and 18 to unidentified receptors. After performing multiple sequence alignments within different biological organisms and antigenicity analysis, we were able to develop antibodies for prawn AMPA ionotropic glutamate receptor 1, metabotropic glutamate receptor 1 and 4, and ionotropic NMDA glutamate receptor subunit 2B, with the expectation that the availability of these antibodies will help broaden knowledge regarding the underlying structural and functional mechanisms involved in prawn behavioral responses to environmental impacts. The Macrobrachium carcinus brain transcriptome can be an important tool for examining changes in many other nervous system molecules as a function of developmental stages, or in response to particular conditions or treatments.
甲壳类动物是水生生态系统的主要组成部分，因此，它们的行为以及身体结构和功能的变化可以作为其周围环境变化的指标，例如与气候变化和人为污染相关的变化。我们使用生物信息学和从头转录组组装方法来鉴定潜在的靶点，以开发针对淡水虾 Macrobrachium spp. 的特定抗体，作为其神经系统功能的标志物。从淡水虾 Macrobrachium carcinus 的脑神经节中提取总 RNA，并使用鳗鱼塘 mRNA Seq 方案进行 Illumina 下一代测序，以构建从头转录组。测序产生了 97,202,662 条序列：47,630,546 对和 1,941,570 条单链。使用 Trinity 进行组装得到了 197,898 个组装的连续体，其中 30,576 个被注释：9,600 个通过同源性，17,197 个通过同源性，3,779 个通过转录家族。我们寻找谷氨酸受体连续体，因为它们在甲壳类动物兴奋性神经传递中起主要作用，发现 138 个与离子型受体相关的连续体，32 个与代谢型受体相关的连续体，18 个与未识别的受体相关的连续体。在对不同生物的多个序列进行比对并进行抗原性分析后，我们能够为对虾 AMPA 离子型谷氨酸受体 1、代谢型谷氨酸受体 1 和 4 以及离子型 NMDA 谷氨酸受体亚基 2B 开发抗体，期望这些抗体的可用性将有助于拓宽对虾对环境影响的行为反应所涉及的潜在结构和功能机制的认识。Macrobrachium carcinus 脑转录组可以成为研究许多其他神经系统分子随发育阶段变化或对特定条件或处理反应的重要工具。
