DOI: 10.5061/dryad.t1g1jwt99
title: Transcriptomes of six Streptanthoid Complex species

description: To increase the number of genomic resources available for species of
 Streptanthoid Complex, we created transcriptomes for six jewelflower
 species. Focal species from six branches were selected to cover a majority
 of the phylogenetic tree. The goal is for these transcriptome sequences to
 aide future studies which look to explore the evolution and adaptation of
 the Streptanthoid Complex as it radiated throughout the California
 Floristic Province.
description: <em>Seed Source</em> The six
 species chosen for transcriptome sequencing were <em>Caulanthus
 anceps, Caulanthus amplexicaulis, Caulanthus inflatus, Streptanthus
 breweri, Streptanthus glandulosus, and Streptanthus tortuosus.</em>
 Seeds for these species were produced in a screenhouse at UC Davis in 2019
 using field collected seeds from 2018 and stored in brown envelopes at
 room temperature. In 2021, these seeds were used to extract RNA under 10
 different tissue and treatment combinations (Table 1).
 Tissue and treatment combinations 
 
 <strong>Combination</strong> 
 <strong>Tissue</strong>
 
 <strong>Treatment</strong> 
 <strong>Timepoints</strong>
 1
 Seed 
 Dry 20 °C 1
 2
 Seed 
 Chilled, Imbibed, 4 °C 
 1 
 3 Leaf
 Normal, 20 °C 
 4 
 4 
 Leaf Cold, 4
 °C 4 
 5 
 Leaf Hot, 40
 °C 4 
 6 
 Leaf Dark,
 20 °C 4 
 7 
 Leaf 
 Drought, 20 °C 
 4 
 8 Root
 Normal, 20 °C 
 1 
 9 
 Flower Normal, 20
 °C 1 
 10 
 Silique 
 Normal, 20 °C 
 1 
 <em>Tissue Collection</em> For
 combination one, dry seeds were removed from their envelopes and
 immediately frozen in liquid nitrogen. Combination two, dry seeds were
 removed from their envelopes, placed on top of germination paper in
 two-inch petri dishes, and imbibed with 3 mL of water. The petri dishes
 were then placed in a 4 °C chamber with constant light for six hours.
 After six hours the seeds were dried to remove excess water and frozen in
 liquid nitrogen. Combinations three through ten were
 collected from young plants. Randomized cones containing a mixture of 50%
 Ron’s Mix and 50% sand were saturated with nutrient water. A small divot
 was then made in each cone where 3-4 seeds were placed before being
 covered with a small amount of the soil mixture. The cones were then
 placed in a rack and covered with plastic wrap to prevent the top layer of
 the soil from drying out. The covered rack was then placed in a growth
 chamber set to 20 °C with a light/dark cycle of 12/12. Two weeks after
 being placed in the growth chamber, the plastic wrap was removed, and the
 recently germinated seedlings were exposed to the air. The seedlings
 remained in the growth chamber with nutrient water being provided every
 other day. Once the seedlings on average had attained approximately 8-10
 true leaves, the seedlings were moved to their treatment conditions and/or
 had their tissue collected. Tissue across multiple replicates of the same
 tissue/treatment combination and different timepoints were pooled and
 frozen. Combination 3 was collected 0, 6, 12, and 18
 hours after lights on. Combination 4 was collected 3, 6, 12, and 24 hours
 after being moved to a 4 °C with a light/dark cycle of 12/12. Combination
 5 was collected .25, .5, 1, and 3 hours after being moved to a 40 °C with
 a light/dark cycle of 12/12. Combination 6 was subjected to a full day
 without light and the following day tissue was collected in the dark 0, 6,
 12, and 18 hours after usual lights on. Combination 7 was subjected to
 five days without watering and on the sixth day tissue was collected in
 the 0, 6, 12, and 18 hours after lights on. Plants for combination 8 were
 removed from their pots, had excess soil washed from their roots, and
 cleaned root tissue collected. Combinations 9 and 10 had their flowers and
 siliques collected once an adequate amount had formed.
 <em>RNA Extraction</em> <em>and
 sequencing</em> Total RNA from 50 mg of each
 sample was extracted using New England Biolab’s Monarch Total RNA Miniprep
 Kit. Total RNA was then quantified, and quality checked on the Qubit 3.
 For each species total RNA from each tissue/treatment was equally pooled
 and 300 ng of total RNA was used with the SMRTbell Express Template Prep
 Kit 2.0 to create a total of 6 SMRTbell libraries. The 6 SMRTbell
 libraries were then sent to the University of California, Davis Genome
 Center for sequencing on the Sequel II. Following sequencing, the
 unaligned BAM files were processed using PacBio’s IsoSeq pipeline
 (“PacificBiosciences/pbbioconda) to create high quality full length
 non-concatemer reads. <em>Transcripts,
 annotation, and orthogroups</em> Transcript
 identification for each species was completed using the isON transcript
 analysis pipeline (Petri and Sahlin, 2023; Sahlin and Medvedev, 2020). The
 pipeline works on PacBio sequence data in two steps, first by clustering
 reads based on sequence similarity and then by generating isoforms out of
 clustered long reads. Following isoform identification, the transcripts
 were then aligned to the UniProt database (The UniProt Consortium, 2023)
 using blastx (Sayers et al., 2022). Annotations were added to the
 transcripts based on their best match. To aide in future investigations,
 orthogroups were generated using orthofinder (Emms and Kelly,
 2019).
description: # Transcriptomes of six Streptanthoid Complex species
 [https://doi.org/10.5061/dryad.t1g1jwt99](https://doi.org/10.5061/dryad.t1g1jwt99) Transcript sequences of *Caulanthus anceps, Caulanthus amplexicaulis, Caulanthus inflatus, Streptanthus breweri, Streptanthus glandulosus, and Streptanthus tortuosus.* Transcripts generated using the isON transcript analysis pipeline. Orthogroup data created using orthofinder. ## Description of the data and file structure Transcriptomes: The 6 transcriptome fasta files[Proceed to Upload](https://datadryad.org/stash/resources/286074/upload) * C. amplexicaulis = C.amplexicaulis_transcriptome.fasta * C. anceps = C.anceps_transcriptome.fasta * C. inflatus = C.inflatus_transcriptome.fasta * S. breweri = S.breweri_transcriptome.fasta * S. glandulosus = S.glandulosus_transcriptome.fasta * S. tortuosus = S.tortuosus_transcriptome.fasta OrthoFinder_Results.tar.gz = The output from orthofinder run using default settings * Citations.txt * Phylogenetic_Hierarchical_Orthogroups * Comparative_Genomics_Statistics * Phylogenetically_Misplaced_Genes * Gene_Duplication_Events * Putative_Xenologs * Gene_Trees * Resolved_Gene_Trees * Log.txt * Single_Copy_Orthologue_Sequences * Orthogroup_Sequences * Species_Tree * Orthogroups * WorkingDirectory * Orthologues ## Sharing/Access information Sequence data can be accessed under **BioProject: PRJNA992064**