本例使用三代组装的玉米NAM群体为例。通过AnchorWave进行全基因组比对，通过gatk进行变异检测。通过tassel进行格式转换。

1.使用AnchorWave进行双序列比对

基因组数据下载链接https://download.maizegdb.org/。NAM群体一共有26个基因组。

anchorwave genoAli -i Zm-B73-REFERENCE-NAM-5.0_Zm00001eb.1.gff3 -as cds.fa -r B73.ref.fa -a B97.sam -ar B73.sam -s Zm-B97-REFERENCE-NAM-1.0.fa -n B97.anchors -o B97.maf -f B97.f.maf -w 38000 -fa3 200000 -B -6 -O1 -8 -E1 -2 -O2 -75 -E2 -1 -IV  >B97.log 2>&1

2.1使用tassel将上步产生的MAF文件转化为GVCF

#download tassel and reference genome.
git clone https://bitbucket.org/tasseladmin/tassel-5-standalone.git
wget https://download.maizegdb.org/Zm-B73-REFERENCE-NAM-5.0/Zm-B73-REFERENCE-NAM-5.0.fa.gz
gunzip Zm-B73-REFERENCE-NAM-5.0.fa.gz/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/B97/B97.maf -sampleName B97_anchorwave -gvcfOutput /home/xuql/copyNAM/B97/B97ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/B97/B97_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML103/CML103.maf -sampleName CML103_anchorwave -gvcfOutput /home/xuql/copyNAM/CML103/CML103ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML103/CML103_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML228/CML228.maf -sampleName CML228_anchorwave -gvcfOutput /home/xuql/copyNAM/CML228/CML228ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML228/CML228_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML247/CML247.maf -sampleName CML247_anchorwave -gvcfOutput /home/xuql/copyNAM/CML247/CML247ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML247/CML247_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML277/CML277.maf -sampleName CML277_anchorwave -gvcfOutput /home/xuql/copyNAM/CML277/CML277ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML277/CML277_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML322/CML322.maf -sampleName CML322_anchorwave -gvcfOutput /home/xuql/copyNAM/CML322/CML322ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML322/CML322_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML333/CML333.maf -sampleName CML333_anchorwave -gvcfOutput /home/xuql/copyNAM/CML333/CML333ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML333/CML333_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML52/CML52.maf -sampleName CML52_anchorwave -gvcfOutput /home/xuql/copyNAM/CML52/CML52ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML52/CML52_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/CML69/CML69.maf -sampleName CML69_anchorwave -gvcfOutput /home/xuql/copyNAM/CML69/CML69ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/CML69/CML69_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/HP301/HP301.maf -sampleName HP301_anchorwave -gvcfOutput /home/xuql/copyNAM/HP301/HP301ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/HP301/HP301_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Il14H/Il14H.maf -sampleName Il14H_anchorwave -gvcfOutput /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Il14H/Il14H_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Ki11/Ki11.maf -sampleName Ki11_anchorwave -gvcfOutput /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Ki11/Ki11_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Ki3/Ki3.maf -sampleName Ki3_anchorwave -gvcfOutput /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Ki3/Ki3_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Ky21/Ky21.maf -sampleName Ky21_anchorwave -gvcfOutput /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Ky21/Ky21_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/M162W/M162W.maf -sampleName M162W_anchorwave -gvcfOutput /home/xuql/copyNAM/M162W/M162WToB73.gvcf -fillGaps false > /home/xuql/copyNAM/M162W/M162W_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/M37W/M37W.maf -sampleName M37W_anchorwave -gvcfOutput /home/xuql/copyNAM/M37W/M37WToB73.gvcf -fillGaps false > /home/xuql/copyNAM/M37W/M37W_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Mo18W/Mo18W.maf -sampleName Mo18W_anchorwave -gvcfOutput /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Mo18W/Mo18W_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Ms71/Ms71.maf -sampleName Ms71_anchorwave -gvcfOutput /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Ms71/Ms71_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/NC350/NC350.maf -sampleName NC350_anchorwave -gvcfOutput /home/xuql/copyNAM/NC350/NC350ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/NC350/NC350_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/NC358/NC358.maf -sampleName NC358_anchorwave -gvcfOutput /home/xuql/copyNAM/NC358/NC358ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/NC358/NC358_outputMafToGVCF.txt/home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Oh43/Oh43.maf -sampleName Oh43_anchorwave -gvcfOutput /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Oh43/Oh43_outputMafToGVCF.txt /home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Oh7B/Oh7B.maf -sampleName Oh7B_anchorwave -gvcfOutput /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Oh7B/Oh7B_outputMafToGVCF.txt /home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/P39/P39.maf -sampleName P39_anchorwave -gvcfOutput /home/xuql/copyNAM/P39/P39ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/P39/P39_outputMafToGVCF.txt /home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Tx303/Tx303.maf -sampleName Tx303_anchorwave -gvcfOutput /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Tx303/Tx303_outputMafToGVCF.txt /home/xuql/tassel-5-standalone/run_pipeline.pl -Xmx100G -debug -MAFToGVCFPlugin -referenceFasta /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -mafFile /home/xuql/copyNAM/Tzi8/Tzi8.maf -sampleName Tzi8_anchorwave -gvcfOutput /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf -fillGaps false > /home/xuql/copyNAM/Tzi8/Tzi8_outputMafToGVCF.txt

2.2对gvcf文件进行压缩和建立索引

对参考基因组，1）更改染色体名，和比对基因组的一样2）建立fai索引3）建立dict索引

sed -i 's/chr//g' Zm-B73-REFERENCE-NAM-5.0.fa
samtools faidx Zm-B73-REFERENCE-NAM-5.0.fa
wget https://github.com/broadinstitute/picard/releases/download/2.26.10/picard.jar
java -jar picard.jar CreateSequenceDictionary R=Zm-B73-REFERENCE-NAM-5.0.fa O=Zm-B73-REFERENCE-NAM-5.0.dict

对gvcf文件：压缩和建立索引，“GATK GenomicsDBImport” 需要这一步。

bgzip -c  /home/xuql/copyNAM/B97/B97ToB73.gvcf > /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML103/CML103ToB73.gvcf > /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML228/CML228ToB73.gvcf > /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML247/CML247ToB73.gvcf > /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML277/CML277ToB73.gvcf > /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML322/CML322ToB73.gvcf > /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML333/CML333ToB73.gvcf > /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML52/CML52ToB73.gvcf > /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/CML69/CML69ToB73.gvcf > /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/HP301/HP301ToB73.gvcf > /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf > /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf > /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf > /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf > /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/M162W/M162WToB73.gvcf > /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/M37W/M37WToB73.gvcf > /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf > /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf > /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/NC350/NC350ToB73.gvcf > /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/NC358/NC358ToB73.gvcf > /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf > /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf > /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/P39/P39ToB73.gvcf > /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf > /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz
bgzip -c  /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf > /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gztabix -p vcf /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz

3.变异检测

3.1由于本步中gatk无法识别大于10M的变异，所以我们通过一下步骤找到>10M的变异并把它删除掉。

#for indel how to sort by python and the part results is shown.
with open('./job.407755.err') as f:lines = f.readlines()
new_lines=[]
for line in lines:if "Set" in line:new_lines.append(line)sorted(new_lines, key=lambda x: int(x.strip().split('=')[-1]),reverse=True)#thoese content output from all chromosome by "gatk LeftAlignAndTrimVariants" and then sorted by python. at last,check the initial output content to identify the super-indel belong to individuals.
['10:56:14.335 INFO  LeftAlignAndTrimVariants - Indel is too long (34461688) at position 9:3695105; skipping that record. Set --max-indel-length >= 34461688\n','10:56:30.429 INFO  LeftAlignAndTrimVariants - Indel is too long (10668738) at position 10:33212598; skipping that record. Set --max-indel-length >= 10668738\n','10:56:28.937 INFO  LeftAlignAndTrimVariants - Indel is too long (9101264) at position 10:14179; skipping that record. Set --max-indel-length >= 9101264\n','10:56:30.038 INFO  LeftAlignAndTrimVariants - Indel is too long (7918835) at position 10:22996027; skipping that record. Set --max-indel-length >= 7918835\n','11:31:49.968 INFO  LeftAlignAndTrimVariants - Indel is too long (7154442) at position 6:16715313; skipping that record. Set --max-indel-length >= 7154442\n',#acquire the line number of super-indel and delete it.
less Oh7BToB73.gvcf | awk '$2=="3695105"{printf("%5d\t%s\n", NR, $0)}' >aaa.txt
less Oh7BToB73.gvcf | sed '32542168d' > aOh7BToB73.gvcf
# compress and index for "gatk GenomicsDBImport"
bgzip -c  /home/xuql/copyNAM/Oh7B/aOh7BToB73.gvcf > /home/xuql/copyNAM/Oh7B/aOh7BToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Oh7B/aOh7BToB73.gvcf.gzless Oh7BToB73.gvcf | awk '$2=="33212598"{printf("%5d\t%s\n", NR, $0)}' >bbb.txt
less Oh7BToB73.gvcf | sed '35176648d' > bOh7BToB73.gvcf
bgzip -c  /home/xuql/copyNAM/Oh7B/bOh7BToB73.gvcf > /home/xuql/copyNAM/Oh7B/bOh7BToB73.gvcf.gz
tabix -p vcf /home/xuql/copyNAM/Oh7B/bOh7BToB73.gvcf.gz

3.2建立GenomicsDBImport

根据实际情况使用内存，对每条染色体都运行这一步，一共10个染色体。

gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 5 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk1 \--genomicsdb-segment-size 1048576000 --genomicsdb-vcf-buffer-size 10000000000 -L 1#Elapsed time: 105.02 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk2 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 2#Elapsed time: 76.68 minutes.  Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk3 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 3#Elapsed time: 70.12 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk4 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 4#Elapsed time: 68.47 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk5 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 5# Elapsed time: 64.29 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk6 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 6#Elapsed time: 53.32 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk7 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 7#Elapsed time: 57.26 minutes.  Runtime.totalMemory()=274877906944gatk --java-options "-Xmx256g -Xms256g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/Oh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk8 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 8#Elapsed time: 52.72 minutes. Runtime.totalMemory()=274877906944gatk --java-options "-Xmx128g -Xms5g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/aOh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk9 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 9#Elapsed time: 54.09 minutes.Runtime.totalMemory()=5368709120gatk --java-options "-Xmx128g -Xms5g" GenomicsDBImport \-V /home/xuql/copyNAM/B97/B97ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML247/CML247ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML333/CML333ToB73.gvcf.gz \-V /home/xuql/copyNAM/HP301/HP301ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki3/Ki3ToB73.gvcf.gz \-V /home/xuql/copyNAM/M37W/M37WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC350/NC350ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh7B/bOh7BToB73.gvcf.gz \-V /home/xuql/copyNAM/Tzi8/Tzi8ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML103/CML103ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML277/CML277ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML52/CML52ToB73.gvcf.gz \-V /home/xuql/copyNAM/Il14H/Il14HToB73.gvcf.gz \-V /home/xuql/copyNAM/Ky21/Ky21ToB73.gvcf.gz \-V /home/xuql/copyNAM/Mo18W/Mo18WToB73.gvcf.gz \-V /home/xuql/copyNAM/NC358/NC358ToB73.gvcf.gz \-V /home/xuql/copyNAM/P39/P39ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML228/CML228ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML322/CML322ToB73.gvcf.gz \-V /home/xuql/copyNAM/CML69/CML69ToB73.gvcf.gz \-V /home/xuql/copyNAM/Ki11/Ki11ToB73.gvcf.gz \-V /home/xuql/copyNAM/M162W/M162WToB73.gvcf.gz \-V /home/xuql/copyNAM/Ms71/Ms71ToB73.gvcf.gz \-V /home/xuql/copyNAM/Oh43/Oh43ToB73.gvcf.gz \-V /home/xuql/copyNAM/Tx303/Tx303ToB73.gvcf.gz \--batch-size 1 \--genomicsdb-workspace-path /home/xuql/copyNAM/NAM_out_gatk10 \--genomicsdb-segment-size 10485760 --genomicsdb-vcf-buffer-size 100000000 -L 10  #Elapsed time: 44.05 minutes.Runtime.totalMemory()=5368709120

3.3变异检测

gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk1 -O /home/xuql/copyNAM/gatk1.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Processed 251973478 total variants in 581.3 minutes. Elapsed time: 581.36 minutes. Runtime.totalMemory()=6694109184gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk2 -O /home/xuql/copyNAM/gatk2.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
# Processed 202340450 total variants in 471.8 minutes.Elapsed time: 472.05 minutes.Runtime.totalMemory()=2147483648gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk3 -O /home/xuql/copyNAM/gatk3.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
# Processed 188148248 total variants in 444.9 minutes.Elapsed time: 445.13 minutes.Runtime.totalMemory()=2147483648gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk4 -O /home/xuql/copyNAM/gatk4.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Processed 188872000 total variants in 432.5 minutes. Elapsed time: 432.67 minutes.Runtime.totalMemory()=2147483648gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk5 -O /home/xuql/copyNAM/gatk5.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Elapsed time: 434.09 minutes.Runtime.totalMemory()=2147483648gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk6 -O /home/xuql/copyNAM/gatk6.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
# Processed 148426585 total variants in 338.4 minutes.Elapsed time: 338.57 minutes. Runtime.totalMemory()=7088373760gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk7 -O /home/xuql/copyNAM/gatk7.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Processed 148023151 total variants in 369.9 minutes.Elapsed time: 370.07 minutes. Runtime.totalMemory()=2147483648gatk --java-options "-Xmx50g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk8 -O /home/xuql/copyNAM/gatk8.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Processed 147858803 total variants in 355.4 minutes.Elapsed time: 355.56 minutes.Runtime.totalMemory()=7214202880#to make sure "GenotypeGVCFs" can implement we delete the indels more than 10M that only included in cheomosome 9 and chromosome 10.gatk --java-options "-Xmx100g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk9 -O /home/xuql/copyNAM/gatk9.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Elapsed time: 320.96 minutes.Runtime.totalMemory()=3724541952gatk --java-options "-Xmx200g" GenotypeGVCFs -R /home/xuql/copyNAM/B73/Zm-B73-REFERENCE-NAM-5.0.fa -stand-call-conf 0 -ploidy 1 -V gendb:///home/xuql/copyNAM/NAM_out_gatk10 -O /home/xuql/copyNAM/gatk10.vcf.gz --cloud-prefetch-buffer 10000 --cloud-index-prefetch-buffer 10000 --genomicsdb-max-alternate-alleles 110 --max-alternate-alleles 100 --gcs-max-retries 1000
#Elapsed time: 291.33 minutes.Runtime.totalMemory()=7079985152

3.4变异标准化

左对齐，修正由于比对策略导致的偏差

/home/ywt/vt/vt normalize gatk1.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk1bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk1bcfvt.vcf.gz -Oz -o gatk1bcftools.vcf.gz
bcftools index -t gatk1bcfvt.vcf.gz
bcftools index -t gatk1bcftools.vcf.gz/home/ywt/vt/vt normalize gatk2.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk2bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk2bcfvt.vcf.gz -Oz -o gatk2bcftools.vcf.gz
bcftools index -t gatk2bcfvt.vcf.gz
bcftools index -t gatk2bcftools.vcf.gz/home/ywt/vt/vt normalize gatk3.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk3bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk3bcfvt.vcf.gz -Oz -o gatk3bcftools.vcf.gz
bcftools index -t gatk3bcfvt.vcf.gz
bcftools index -t gatk3bcftools.vcf.gz/home/ywt/vt/vt normalize gatk4.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk4bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk4bcfvt.vcf.gz -Oz -o gatk4bcftools.vcf.gz
bcftools index -t gatk4bcfvt.vcf.gz
bcftools index -t gatk4bcftools.vcf.gz/home/ywt/vt/vt normalize gatk5.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk5bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk5bcfvt.vcf.gz -Oz -o gatk5bcftools.vcf.gz
bcftools index -t gatk5bcfvt.vcf.gz
bcftools index -t gatk5bcftools.vcf.gz/home/ywt/vt/vt normalize gatk6.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk6bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk6bcfvt.vcf.gz -Oz -o gatk6bcftools.vcf.gz
bcftools index -t gatk6bcfvt.vcf.gz
bcftools index -t gatk6bcftools.vcf.gz/home/ywt/vt/vt normalize gatk7.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk7bcfvt.vcf.gz
bcftools norm  -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk7bcfvt.vcf.gz -Oz -o gatk7bcftools.vcf.gz
bcftools index -t gatk7bcfvt.vcf.gz
bcftools index -t gatk7bcftools.vcf.gzbcftools index -t gatk8.vcf.gz
/home/ywt/vt/vt normalize gatk8.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk8bcfvt.vcf.gz
bcftools norm -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk8bcfvt.vcf.gz -Oz -o gatk8bcftools.vcf.gz
bcftools index -t gatk8bcfvt.vcf.gz
bcftools index -t gatk8bcftools.vcf.gz/home/ywt/vt/vt normalize gatk9.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk9bcfvt.vcf.gz
bcftools norm -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk9bcfvt.vcf.gz -Oz -o gatk9bcftools.vcf.gz
bcftools index -t gatk9bcfvt.vcf.gz
bcftools index -t gatk9bcftools.vcf.gz/home/ywt/vt/vt normalize gatk10.vcf.gz -r /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -o gatk10bcfvt.vcf.gz
bcftools norm -f /media/ywt/14T1/NAManchorwave/B73/Zm-B73-REFERENCE-NAM-5.0.fa -m +both gatk10bcfvt.vcf.gz -Oz -o gatk10bcftools.vcf.gz
bcftools index -t gatk10bcfvt.vcf.gz
bcftools index -t gatk10bcftools.vcf.gz

4使用IGV可视化结果

sed -r -i 's/[0-9]+H//g' B97.sam &
samtools view -O CRAM --threads 80 --reference Zm-B73-REFERENCE-NAM-5.0.fa B97.sam | samtools sort --threads 30 -O CRAM - > B97.cram
samtools index B97.cram

三代基因组全基因组的变异检测相关推荐

基于三代测序数据的结构变异检测，PBHoney方法解读
本文来自"生信算法"公众号. 基因变异普遍存在于同一物种内的不同个体中,如人与人之间的基因组是不完全一样的(即是多态的),彼此之间都存在着一些差异,即使是和父母或是兄弟姐妹之间去比 ...
推荐一款高引超6000次的全基因组/全外显子组变异注释工具
SnpEff是韦恩州立大学Douglas M. Ruden团队于2012年发表的一款变异注释工具.到现在已近10年历史,持续更新,已至5.0版,总引用6044次. SnpEff的优点基于Java环境 ...
[原]基因组变异检测概述
考虑到cnblog不适合基因组领域这种类型的文章,进过多番折腾,终于用jekyll+github搭了个独立博客www.huangshujia.me,现在博客已经搬迁! 首先,在开始之前我觉得有必要稍微 ...
基因组变异检测概述（SNP、InDel、SV）
首先,在开始之前我觉得有必要稍微科普缓冲一下,以便不使得不熟悉生物信息或基因组的客官们疑惑.O(∩_∩)O! 1.基因组:每个人都有一个基因组,这里的"基因组"并不只是" ...
全基因组关联分析中上位性检测算法的研究
全基因组关联分析中上位性检测算法的研究前言这个项目主要是分享一些全基因组关联分析中上位性检测算法的研究经验,算是,怎么入门,写这么个东西,一是做总结,二是咱实验室估计以后还会有做这个方向的,备着吧 ...
signature=d363d26bda212f777fef81d270ecd42b,基于DNA-pooling全基因组重测序初步筛查CAD易感基因变异位点...
摘要: 目的:应用DNApooling全基因组重测序技术初步筛查CAD易感基因变异位点并进行功能分析.方法:分别收集CAD病例组血液样本和正常对照组血液样本各100例.提取病例组和对照组DNA,制作D ...
基因组大数据变异检测算法的并行优化
基因组大数据变异检测算法的并行优化基因组大数据变异检测算法的并行优化崔英博1, 黄春1, 唐滔1, 杨灿群1, 廖湘科1, 彭绍亮2,3 1 国防科技大学计算机学院,湖南长沙 410073 2 ...
基于全基因组测序数据鉴定结构变异的四大类方法总结
不同类型的基因组变异示意图(图片来源:labspaces) 上次给大家总结介绍了基因组单核苷酸多态性(single nucleotide polymorphism,SNP)的鉴定方法,今天给大家介绍结 ...
全基因DNA甲基化检测“金标准”--- 改进版简化基因组甲基化测序
亚硫酸盐转化被誉为DNA甲基化(5mC)检测的"金标准",非甲基化胞嘧啶C在亚硫酸盐转化过程中转变为尿嘧啶U,进而通过PCR扩增转变为胸腺嘧啶T,甲基化mC维持不变,通过高通量测序 ...

三代基因组全基因组的变异检测