RNA-seq of larval stages around molting and metamorphosis

P. ornatus larvae were cultured from wild-caught broodstock at IMAS, under proprietary culture conditions. Approximately 500 stage 9 and 10 phyllosoma were transferred from a mass rearing tank and distributed between two 110 L Kriesel tanks with flow-through open circulation. Larvae were sampled for RNA-extraction at 12 defined points throughout development. Phyllosoma were harvested at 4 days post-molt in stages 10.1, 10.2, 11.1 and 11.2 (where n.x denotes stage n instar x) - the latter of which culminates in metamorphosis. In this final stage, additional samples were taken at 6 and 8 days post-molt, and at approximately 10 days when the onset of gut-retraction was visible. Further samples were taken from the post-molt, H-phase and pigmented puerulus and then in the juvenile phase at 0 and 4 days post-molt. RNA was extracted from whole larvae (n=3) for sequencing.

P. ornatus幼体由IMAS的野生捕获亲体繁育，采用专有养殖条件培育。约500只第9、10期叶状幼体（phyllosoma）从批量养殖池转移至两个配备开放式流水循环系统的110升Kriesel养殖槽中分养。研究人员在幼体整个发育周期的12个预设时间点取样，用于RNA提取。研究人员于第10.1、10.2、11.1及11.2期叶状幼体蜕壳后4天采集样本（其中n.x代表第n龄期的第x亚期），第11.2期为变态发育的终末阶段。在该终末阶段，研究人员额外于蜕壳后6天、8天以及蜕壳后约10天（此时可见肠道收缩起始）进行取样。后续还分别采集了蜕壳后阶段、H期以及着色幼体（puerulus）的样本，随后在幼体阶段于蜕壳后0天和4天取样。最终从全部幼体中提取RNA（每组生物学重复数为3）用于测序。