The full-length Quillaja brasiliensis (Quillajaceae) reference transcriptome

Introduction

In this study, we used the PacBio Iso-Seq technology to uncover the full-length transcriptome of Quillaja brasiliensis (Quillajaceae), a tree species native to Brazil and of great importance for the extraction of bioactive saponins. Establishing a full-length transcriptome is essential for understanding the metabolic pathways leading to saponin biosynthesis in Q. brasiliensis and should set the stage for upcoming investigations.

Sample obtention and processing

We collected seeds of Quillaja brasiliensis (A.St.-Hil. & Tul.) Mart. (Quillajaceae) from the city of Canguçú (Rio Grande do Sul, Brazil) in March/2018. After in vitro germination, we explanted the seedlings and submitted them to callogenesis or transferred them to the grow room at 25°C under a 16h/day photoperiod. We detached leaves from 2.8-year-old individuals for the experiments hereby presented. After leaf cleanup, we detached approximately 15 leaves and exposed them to ultraviolet radiation (UV-C germicide lamp, ʎ maximum 254 nm) or white light (control) (Table 1). After incubation, we froze the plant material in liquid nitrogen and stored it at -80 °C until the next processing step. We obtained calluses from in vitro germinated explants and submitted them to cellular suspension induction. We maintained the cell cultures in MS medium supplemented with naphthaleneacetic acid (5 mg/L) and kinetin (0.1 mg/L) in the absence of light, using 250 mL culture flasks under the agitation of 120 RPM. After establishing the growth profile, we collected three flasks for each growth phase of the liquid cell cultures: lag phase (three days), log phase (seven days), and stationary phase (21 days). For the sample collection, cells were filtered using the Büchner funnel, washed with distilled water, and flash-frozen in liquid nitrogen. We stored the collected material at -80 °C until the next processing step. We extracted RNA samples from different tissues and growth conditions using the cetyltrimethylammonium bromide method (CTAB) and performed their purification using the RNeasy MinElute Cleanup Kit (QIAGEN, Hilden, Germany). We quality-controlled the samples using fluorometric and spectrophotometric methods before submitting them to sequencing by Novogene (Beijing, People's Republic of China). We made an equimolar pool of the five sample types for sequencing using the Iso-Seq strategy (PacBio, Menlo Park, US) in circular consensus sequencing mode for the obtention of long reads. The raw reads are available at the European Nucleotide Archive (ENA) under accession PRJEB58985.

Sample

Sample source

Treatment or Growth phase

Leaf UV light

Leaf

Ultraviolet light treatment.

Leaf white light

White light treatment

Cell suspension lag

Cell suspension

Lag phase (three days)

Cell suspension log

Log phase (seven days)

Cell suspension stationary

Stationary phase (21 days)

Table 1. Brief description of pooled sequenced samples. 

Zenodo repository content

This repository stores the full-length transcriptome FASTA file obtained after running the IsoSeq v3 pipeline, followed by one round of polishing by LoRDEC and transcript-collapsing by Cogent/Cupcake-ToFU (collapsed_isoforms.fa). We also performed coding sequence prediction and functional annotation using CodAn and TRAPID 2.0 (PLAZA 4.5 Dicots), respectively (transcriptome_functional_characterization.tsv). A brief description of column names for the functional characterization table is also available here.