三代测序长reads中准确和稳定的识别二倍体基因组结构变异（SV）

本文链接：https://blog.csdn.net/LittleComputerRobot/article/details/144848379

VolcanoSV

VolcanoSV能够从单分子长reads中准确和稳定的识别二倍体基因组中的结构变异（SV），拥有单染色体模式和WGS模式两种运行模式。

github: https://www.github-zh.com/projects/623191841-volcanosv#single-chromosome-mode

详细用法参考： https://www.github-zh.com/projects/623191841-volcanosv#single-chromosome-mode

1. 下载安装

# 克隆
git clone --recurse-submodules https://github.com/maiziezhoulab/VolcanoSV.git
cd VolcanoSV
sh Install.sh


# 创建conda环境, python版本3.8.3
conda env create -f VolcanoSV/requirement.yaml
conda activate volcanosv

# 设置VolcanoSV所在目录，方便后续脚本调用
path_to_volcanosv=/path/to/VolcanoSV

VolcanoSV

2. 数据下载

对于单染色体模式，提供了用于Hifi、BamHI和ONT数据的chr10 BAM、contigs和VCF测试文件。可以从Zenodo网站下载： https://zenodo.org/records/10520476

针对输入的BAM数据或ONT数据的结果，则只需将输入BAM文件和参数“dtype”更改为相应的（BAM/ONT）数据类型即可。

# lunix命令行下载， 示例数据使用的hg19参考基因组
wget https://cf.10xgenomics.com/supp/genome/refdata-hg19-2.1.0.tar.gz
tar -xzvf refdata-hg19-2.1.0.tar.gz

3. 单染色体模式

3.1 单染色体模式VolcanoSV组装（VolcanoSV-asm）

VolcanoSV组装流水线被设计为通过染色体运行。我们将多个最先进的汇编器集成到管道中，包括hifiasm，Flye，wtdbg 2，miniasm，Shasta，NextDenovo和Hicanu/Canu。用户可以根据自己的需要选择合适的汇编器。主脚本是${path_to_volcanosv}/bin/VolcanoSV-asm/volcanosv-asm.py。

# 对高保真数据使用hifiasm
# 默认情况下，VolcanoSV对Hifi数据使用hifiasm，对ONT和ONT数据使用Flye
python ${path_to_volcanosv}/bin/VolcanoSV-asm/volcanosv-asm.py \
-bam Hifi_L2_hg19_minimap2_chr10.bam \
-o volcanosv_asm_output \
-ref refdata-hg19-2.1.0/fasta/genome.fa \
-t 10 \
-chr 10 \
-dtype Hifi \
-px Hifi_L2 \
-asm hifiasm

参数说明：
–bam_file INBAM, -bam INBAM, could be either wgs bam or single-chromosome bam file
–output_dir output_dir, -o output_dir
–reference REFERENCE, -ref REFERENCE
–n_thread N_THREAD, -t N_THREAD
–chrnum {1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22}, -chr {1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22}
–assembler {wtdbg2,canu,miniasm,shasta,nextdenovo,hifiasm,hicanu,flye}, -asm {wtdbg2,canu,miniasm,shasta,nextdenovo,hifiasm,hicanu,flye}
optional; if not set, VolcanoSV use hifiasm for Hifi data and flye for CLR and ONT data by default.
–data_type {CLR,ONT,Hifi}, -dtype {CLR,ONT,Hifi}
–pacbio_subtype {rs,sq}, -pb {rs,sq}
must provide when using wtdbg2 on CLR data (default: None)
–shasta_ont_config {Nanopore-OldGuppy-Sep2020}, -shacon {Nanopore-OldGuppy-Sep2020}
–prefix PREFIX, -px PREFIX

3.2 单染色体模式大插入缺失检测（VolcanoSV-vc）

主脚本${path_to_volcanosv}/bin/VolcanoSV-vc/Large_INDEL/volcanosv-vc-large-indel.py

输入目录应该是volcanoSV-asm的输出目录。脚本兼容单染色体模式或wgs模式：当提供参数“chrnum”时，它将在单染色体模式下执行，否则，它将假设input_dir包含chr1-chr 22并在wgs模式下执行。请注意前缀prefix应该与volcanosv-asm中的设置保持一致。运行上述代码后，您将在<ouput_folder>/volcanosv_large_indel.vcf中输出VCF。

python ${path_to_volcanosv}/bin/VolcanoSV-vc/Large_INDEL/volcanosv-vc-large-indel.py \
-i volcanosv_asm_output/ \
-o volcanosv_large_indel_output/ \
-dtype Hifi \
-bam Hifi_L2_hg19_minimap2_chr10.bam \
-ref refdata-hg19-2.1.0/fasta/genome.fa \
-chr 10 -t 10 \
-px Hifi_L2

参数说明：
–input_dir INPUT_DIR, -i INPUT_DIR
–output_dir OUTPUT_DIR, -o OUTPUT_DIR
–data_type DATA_TYPE, -dtype DATA_TYPE
Hifi;CLR;ONT
–bam_file RBAM_FILE, -bam RBAM_FILE
reads bam file for reads signature extraction
–reference REFERENCE, -ref REFERENCE
wgs reference file
–chrnum {1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22}, -chr {1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22}
–n_thread N_THREAD, -t N_THREAD
–n_thread_align N_THREAD_ALIGN, -ta N_THREAD_ALIGN
–mem_per_thread MEM_PER_THREAD, -mempt MEM_PER_THREAD
Set maximum memory per thread for alignment; suffix K/M/G recognized; default = 768M
–prefix PREFIX, -px PREFIX

3.3 单染色体模式小插入缺失检测（VolcanoSV-vc）

主脚本是${path_to_volcanosv}/bin/VolcanoSV-vc/Small_INDEL/volcanosv-vc-small-indel.py。将在volcanosv_small_indel/Hifi_L2_volcanosv_small_indel.vcf中输出VCV。

python ${path_to_volcanosv}/bin/VolcanoSV-vc/Small_INDEL/volcanosv-vc-small-indel.py \
-i volcanosv_asm_output/ \
-o volcanosv_small_indel \
-bam Hifi_L2_hg19_minimap2_chr10.bam \
-ref refdata-hg19-2.1.0/fasta/genome.fa \
-r chr10 \
-t 30 \
-px Hifi_L2

参数说明：
–input_dir INPUT_DIR, -i INPUT_DIR
–bam_file READ_BAM, -bam READ_BAM
–output_dir OUTPUT_DIR, -o OUTPUT_DIR
–reference REFERENCE, -ref REFERENCE
–bedfile BEDFILE, -bed BEDFILE
optional; a high confidence bed file (default: None)
–region REGION, -r REGION
optional; exmaple: chr21:2000000-2100000 (default: None)
–n_thread N_THREAD, -t N_THREAD
–kmer_size KMER_SIZE, -k KMER_SIZE
–ratio RATIO, -rt RATIO
maximum bad kmer ratio (default: 0.3)
–min_support MIN_SUPPORT, -ms MIN_SUPPORT
maximum support for bad kmer (default: 5)
–prefix PREFIX, -px PREFIX