SNS

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CNV calling on single-cell genome sequencing data[edit]

CSHL SNS caller[edit]

  • The method was developed by CSHL Baslan et al. Nat Protoc, 2012.
  • A Perl script fastq2cnv.pl was written to implement this method in both Triton and Genome-miner.
  • To run this script, one needs a .info file per sample. The file should contain the server name (the first line, either triton or genome-miner), location and other information of the sequencing data. Here is an example of a .info file.
  • On genome-miner, the job can be started with the following command:
  nohup PATH_NAME/fastq2cnv.pl  SAMPLE_NAME.info > SAMPLE_NAME_SNS.log &
  • On Triton, a job file needs to be created for each sample. And the job can be submitted by qsub. Here is an example of .job file.
  qsub SAMPLE_NAME.job
  • Once the job is completed, transfer the SAMPLE_NAME.varbin50k.out to your local computer (you can also run R in genome-miner, but not triton).
  • Edit two R scripts by replacing the sample name with the actual one you are using.
  SAMPLE_NAME.cbs.r
  SAMPLE_NAME.copynumber.r
  • Under R, run the following two commands (make sure both the varbin50k.out file and the R scripts are in the current directory).
  source("SAMPLE_NAME.cbs.r");
  source("SAMPLE_NAME.copynumber.r");
  • You will find two dozens of plots in the Postscript format, plus a number of other text files. Postscript files can be converted into pdf files using Acrobat Distiller, or ps2pdf in Unix/Linux.
   SAMPLE_NAME.hg19.50k.k50.varbin.data.copynumber.
   SAMPLE_NAME.hg19.50k.k50.nobad.varbin.data
   SAMPLE_NAME.hg19.50k.k50.nobad.varbin.short
   SAMPLE_NAME.hg19.50k.k50.varbin.data
   SAMPLE_NAME.hg19.50k.k50.varbin.short

BIC-Seq[edit]

  • This method was developed by Peter Park's group at Harvard. Xi et al. PNAS 2011
  • It starts with mapped bam files after removal of clonal reads.
  • Step 1: generate read count files from the bam file (replace DIR with a directory name to write all the .seq files).
 /home/kunzhang/softwares/BICseq/PERL/BICseq_1.1.2/SAMgetUnique/samtools-0.1.7a_getUnique-0.1.1 view -U BWA,DIR/,N,N SAMPLE_NAME.bam
 Repeat the same procedure for each bam files, use one output directory per sample
  • Step 2: create a tab-delimited config file as the following:
 chrom   case    control
 chr1    case/chr1.seq   control/chr1.seq
 chr2    case/chr2.seq   control/chr2.seq
  • Step 3: run BIC-seq.pl:
 /home/kunzhang/softwares/BICseq/PERL/BICseq_1.1.2/BIC-seq/BIC-seq.pl [options] <ConfigFile> <OutputDir> <Description>
 Options:
       --help
       --lambda=<float>: default 2
       --bin_size=<int>: default 100
       --multiplicity=<float>: default 2
       --window=<int>: the window for removing the outliers; default 200
       --f=<float>: expected number of type I errors in the merging process; An alternative way to specify lambda
       --B=<int>: number of permutations for FDR estimate; default 0
       --paired: if specified the data is treated as paired-end data
       --I=<Insert,SDofInsert>: specify the insert size and standard deviation of insert size. Default <200,20>
 <Outputdir>: the name of the directory to write the output (this cannot be an existing directory).
 <Description>: used to name the segmentation results under <OutputDir>
  • For detail, see /home/kunzhang/softwares/BICseq/PERL/BICseq_1.1.2/BIC-seq/README