Illumina GA: Difference between revisions

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m (New page: ==Illumina sequencing== ===Sample information=== *Each sample must have the following information: **Sample ID: for sample tracking, please adopt a consistent naming scheme. A name should ...)
 
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==Illumina sequencing==
*[ftp://ftp.illumina.com Illumina FTP Server]
===Sample information===
*login: FAS_Public
*Each sample must have the following information:
*pass: illumina
**Sample ID: for sample tracking, please adopt a consistent naming scheme. A name should have the following fields separated by hyphens "-". Each field should not contain any hyphen. Use underscore "_" as a separator if necessary. The sample ID is often too long to write on the tube. It's okay to use a shorter name on the tube, but there has to be a look-up table on the wiki page to map a short name to the Sample ID. The full-length IDs have to be used in the GA tracking form. 
 
#Initial of the person who prepared the sample.
 
#Methods used for the sample processing, such as Exome, BisSeq, MDA etc.
*[[Illumina_GA/Oligo_info|Common primers used in Solexa sequencing]]'''
#Names of the starting materials, such as CV_iPS, PGP1 etc.
 
#Batch ID or date of the library preparation.
*[[Illumina_GA/Sample_info|Informatics]]'''
#Additional information.
 
  Here is an example:
*[[Illumina_GA/Run_info|Checklist]]'''
  KZ-Exome-NA12878-Oct16-EL140.3
 
***For external libraries, please use the following fields:
*[[Illumina_GA/Sequencing_Guide_info|Experienced User Card]]'''
#Institution name;
 
#The PI's initial;
*[[Illumina_GA/Repair|PE_Module]]'''
#Label on the tube;
 
#Date of receipt.
*[http://genome-tech.ucsd.edu/LabNotes/upload/large_files/GenomeStudioV2010.1.exe GenomeStudio]:
**PAGE gel image with Invitrogen LowMass ladder as the reference.
**DNA sequencing module: 2863-5528-3159-7360
**Record of QPCR quantification.
**RNA Sequencing module: 3568-8622-6540-4466
===Workflow for post-run processing===
**ChIP Sequencing module: 7542-2534-0192-4527
*The data for each run is stored in the folder /media/NewReads/FreshReads of genome-miner.ucsd.edu
**Genotyping module: 0219-8548-3780-7356
*The run root folder name looks like: 091209_HL029 or 091218_AL006.
 
*The SCS software reports sequences in the qseq format by default.
*Barcode space:
*The .qseq files can be found in the [Run Root Folder]/Data/Intensities/BaseCalls folder, such as
**Alice    Indx13-24
    /media/NewReads/FreshReads/091209_HL029/Data/Intensities/BaseCalls
**Eric     Indx09-32,49-72
*Most downstream processing requires the sequences in the fastq format, one file per lane. There are two options to obtain fastq files:
**Jeff      Indx25-48
**Fistly, create a new folder in the SeqStore directory:
**Sam      Indx49-72
    mkdir /media/SeqStore2/AL006
**Rui      Indx73-96
**If you trust the basecaller of the SCS software, then you may simply use a format converter to generate fastq files:
**Noi      Indx1-96
    cd /media/NewReads/FreshReads/091209_HL029/Data/Intensities/BaseCalls
    cat s_1_?_????_qseq.txt | /usr/local/bin/qseq2fastq.pl > /media/SeqStore2/AL006/s_1_sequence.txt &
    cat s_2_?_????_qseq.txt | /usr/local/bin/qseq2fastq.pl > /media/SeqStore2/AL006/s_2_sequence.txt &
    .
    .
    .
    cat s_8_?_????_qseq.txt | /usr/local/bin/qseq2fastq.pl > /media/SeqStore2/AL006/s_8_sequence.txt &
**In many cases, we need to repeat the base calling using the GA Pipeline because some of the lanes either contain libraries that have biased base composition (bisulfite converted libraries) or the libraries were overloaded. In such cases, we need to us one of the "good lane" to determine the phasing for the base caller. This requires running Bustart and Gerald in the genome-miner. Note that in the second command line you have to set the control lane to the best lane of the flowcell.
    cd /media/NewReads/FreshReads/091209_AL006
    /usr/local/GAPipeline-1.5.1/bin/bustard.py Data/Intensities/ --CIF --control-lane=4 --GERALD=../config.txt -make
    cd Data/Intensities/Bustard1.5.1_21-12-2009_root  #Note that the name of Bustard directory contains the date the previous command was run.
    nohup make recursive -j 8 &
    # this will take a few hours
    # when the base calling is completed, you may find the eight fastq files in the following folders:
     cd /media/NewReads/FreshReads/091218_AL006/Data/Intensities/Bustard1.5.1_21-12-2009_root/GERALD_21-12-2009_root
    cp s_?_sequence.txt > /media/SeqStore2/AL006
    # A summary of base calling is in the file Summary.htm.

Latest revision as of 22:59, 14 October 2014


  • GenomeStudio:
    • DNA sequencing module: 2863-5528-3159-7360
    • RNA Sequencing module: 3568-8622-6540-4466
    • ChIP Sequencing module: 7542-2534-0192-4527
    • Genotyping module: 0219-8548-3780-7356
  • Barcode space:
    • Alice Indx13-24
    • Eric Indx09-32,49-72
    • Jeff Indx25-48
    • Sam Indx49-72
    • Rui Indx73-96
    • Noi Indx1-96