Phragmites-australis-transcriptome-optimal-assembly

Workflow and scripts used for the optimal assembly of the Phragmites australis transcriptome.

Introduction

Multiple types of tissue were used to construct forty-nine P. australis transcriptomes via different assembly tools and multiple parameter settings. The optimal transcriptome for functional annotation and downstream analyses was selected among these transcriptomes by comprehensive assessments. For a total of 422,589 transcripts assembled in this transcriptome, 319,046 transcripts (75.5%) have at least one functional annotation. Within the transcriptome, 1,495 transcripts showing tissue-specific expression pattern, 10,828 putative transcription factors, and 72,165 simple sequence repeats markers were further identified. With this optimal transcriptome and all relative information from downstream analyses, foundations for future studies on the mechanisms underlying the invasiveness of non-native P. australis were laid.

Data

1. P. australis was sampled by the plant phenotypic character from an open field in urban Detrot, MI, US, including four tissues data: leaf, shoot meristem, rhizome and inflorescence. A total of 264 million reads of 100bp Illumina paired end RNA-seq data were obtained. PRJNA762483
2. Setaria italica v2.2 from Phytozome
3. P. australis leaf culm and rhizome tip tissue transcriptome sequence reads from PRJNA314710
4. BUSCO library: poales_odb10 and embryophyta_odb10
5. Uniprot Swiss-Prot (ftp://ftp.uniprot.org/pub/databases/uniprot/current_release/knowledgebase/complete/uniprot_sprot.fasta.gz)
6. Pfam (ftp://ftp.ebi.ac.uk/pub/databases/Pfam/current_release/Pfam-A.hmm.gz)
7. COG (clusters of orthologous genes database)
8. NR (RefSeq non-redundant proteins database) (ftp://ftp.ncbi.nlm.nih.gov/blast/db/FASTA/nr.gz)
9. S. italica transcription factors list from PlantTFDB v5.0

Software

skewer-0.22, Trinity-2.8.4, FastQC v0.11.8, STAR-2.5.2b, shannon-0.02, transabyss-2.0.1, SOAPdenovo-Trans-v1.03, GapCloser-v1.12, bowtie2-2.3.0, CD-HIT-4.8.1, EvidentialGene17dec14, kallisto-0.46.1, ncbi-blast-2.8.1+, DETONATE-1.11, hmmer-3.3, BUSCO 4.0.5, rnaQUAST-2.0.0, q30-master, TransDecoder-v5.5.0, Trinotate-v3.2.1, sqlite-snapshot-202101010144, signalp-4.1g, tmhmm-2.0c, DIAMOND v2.0.6, rsem-1.2.19, Venny, WEGO 2.0, GhostKOALA, MISA v2.1

Data Analyses

1. Before assembly

2. De novo transcriptome assembly using different tools and parameters

3. EvidentialGene pipeline treament

4. Transcriptome quality assessments

5. Transcriptome annotation

6. DETs and GO enrichment analyses

7. SSRs and TFs identification

Results for future study reference

Include all seven compared transcriptomes, predicted peptide sequence file, annotation file and results from downstream analyses

Citation

Feng Tao, Chuanzhu Fan, Yimin Liu, Subashini Sivakumar, Kurt P. Kowalski, Edward M. Golenberg. Optimization and Application of non-native Phragmites australis Transcriptome Assemblies. In review.