This archive includes scripts used to do de novo genome assembly of Sophora Flavescens (苦参, Ku-shen). The sequencing data used to do genome assembly including following datasets:
- Nanopore WGS sequencing: ~100x
- Illumina WGS sequencing: ~100x
- HiC WGS sequencing: ~100x
- Transcriptome sequencing: from leaf, stem and root
The folder structure of this archive is as follows:
./KS_WGS_scripts/
├── [May 18 14:09] 01_genome_survey
├── [May 18 11:25] 02_genome_assembly
│ ├── [May 18 14:02] 01_QC
│ ├── [May 18 14:02] 02_1_raven_raw_all
│ ├── [May 18 14:02] 02_2_raven_raw_N50
│ ├── [May 18 14:03] 03_1_smartdenovo_raw_all
│ ├── [May 18 14:03] 03_2_smartdenovo_raw_N50
│ ├── [May 18 14:03] 04_1_wtdbg2_raw_all
│ ├── [May 18 14:03] 04_2_wtdbg2_raw_N50
│ ├── [May 18 14:03] 05_1_flye_raw_all
│ ├── [May 18 14:04] 05_2_flye_raw_N50
│ ├── [May 18 14:04] 06_1_raven_canu_ec
│ ├── [May 18 14:04] 07_1_smartdenovo_canu_ec
│ ├── [May 18 14:04] 08_1_wtdbg2_canu_ec
│ ├── [May 18 14:04] 09_1_flye_canu_ec
│ ├── [May 18 14:04] 10_1_raven_fmlrc_N50
│ ├── [May 18 14:04] 11_1_smartdenovo_fmlrc_N50
│ ├── [May 18 14:04] 12_1_wtdbg2_fmlrc_N50
│ ├── [May 18 14:05] 13_1_flye_fmlrc_N50
│ ├── [May 18 14:05] e1_canu_ec
│ ├── [May 18 14:05] e2_fmlrc_ec
│ ├── [May 18 14:05] t1_1_purge_haplotigs
│ ├── [May 18 14:05] t1_2_HiC_scaffolding
│ └── [May 18 14:06] t1_canu_assembly
├── [May 18 13:21] 03_genome_annotation
│ ├── [May 18 14:06] 01_gene_annotation
│ ├── [May 18 14:08] 02_repeat_annotation
│ └── [May 18 14:08] 03_transcriptome
└── [Jul 11 2022] 04_phylogenomics
├── [May 18 14:08] 01_orthofinder
├── [May 18 14:07] 02_gene_expansion
└── [May 18 14:07] 03_WGD
33 directories
There are four top level folders, including scripts used to perform four different parts of the analysis in the project.
The folder 01_genome_survey includes scripts used to do genome survey analysis using illumina WGS sequencing dataset.
This folder includes scritps to do de novo genome assembly. To obtain a high-quality reference genome, we used 17 different assembly strategies (see following table). The initial strategy was to use the CANU-only (v2.0) pipeline. After the error-correction of Nanopore reads using the CANU correct module, we used the CANU trim module to remove low quality regions in error-corrected reads. The genome was then assembled using the CANU assemble module. In addition, we also tried four other assemblers, including Raven (v1.1.10), SMARTdenovo (v1.0), wtdbg2 (v1.1) and Flye (v2.7.1) on four different input datasets respectively. The first input dataset includes all nanopore raw reads (named as raw_all). The second input dataset is a subset of the first dataset, including only raw reads longer than the N50 of all raw reads (named as raw_N50). The third input dataset includes error-corrected Nanopore reads using CANU (named as canu_ec). And the fourth input dataset includes error-corrected reads longer than the N50 of all error-corrected reads using FMLRC (v1.0.0) (fmlrc_N50).
| Assembler | Input | Number_of_reads | N50_of_reads | Depth |
|---|---|---|---|---|
| raven | raw_all | 1,968,671,916 | 25.4 | 105.9x |
| smartdenovo | raw_all | 3,104,086,840 | 25.4 | 105.9x |
| wtdbg2 | raw_all | 2,785,948,512 | 25.4 | 105.9x |
| flye | raw_all | 2,785,242,597 | 25.4 | 105.9x |
| raven | raw_N50 | 1,458,531,649 | 37.8 | 52.9x |
| smartdenovo | raw_N50 | 2,173,979,684 | 37.8 | 52.9x |
| wtdbg2 | raw_N50 | 2,790,423,511 | 37.8 | 52.9x |
| flye | raw_N50 | 2,852,564,288 | 37.8 | 52.9x |
| raven | fmlrc_N50 | 390,557,486 | 37.9 | 52.9x |
| smartdenovo | fmlrc_N50 | 2,183,176,129 | 37.9 | 52.9x |
| wtdbg2 | fmlrc_N50 | 4,537,814,869 | 37.9 | 52.9x |
| flye | fmlrc_N50 | 2,307,728,032 | 37.9 | 52.9x |
| raven | canu_ec | 511,724,891 | 20 | 94.2x |
| smartdenovo | canu_ec | 2,520,480,671 | 20 | 94.2x |
| wtdbg2 | canu_ec | 2,786,293,469 | 20 | 94.2x |
| flye | canu_ec | 1,904,306,272 | 20 | 94.2x |
| canu | canu_ec | 3,636,323,899 | 20 | 94.2x |
Haplotigs were then purged from contigs (subfolder t1_1_purge_haplotigs), and then contigs with haplotigs removed were further scaffolded using HiC WGS sequencing dataset. Scripts used to do scaffolding are stored in subfolder t1_2_HiC_scaffolding.
This folder includes scripts to do gene annotation (subfolder 01_gene_annotation), repeat annotation (subfolder 02_repeat_annotation) and transcriptome analysis (subfolder 03_transcriptome).
This folder includes scripts to do phylogenomics for the assembled genome, including:
- 01_orthofinder: Orthologs identification
- 02_gene_expansion: Gene expansion and contraction analysis
- 03_WGD: Whole genome duplication analysis