Athurva Gore/LabNotes/2011-9-23: Difference between revisions

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* Created oligos using script: [[File:probe2padlockCpgLibSept11_fixed09232011.txt]]
* Created oligos using script: [[File:probe2padlockCpgLibSept11_fixed09232011.txt]]
* Final oligos: [[File:NM_LC_Probes_Filtered_09232011.txt.seq]]
* Final oligos: [[File:NM_LC_Probes_Filtered_09232011.txt.seq]]
=Low-input Exomes=
* Alice used small numbers of Lymphocytes to perform Exome Capture from Nextera libraries
** Used suppressor oligonucleotides for Nextera adaptor sequences to hopefully increase specificity of capture
** Used very few cells in each step
* Need to generate statistics for each library (D, E, F)
** Currently doing this blind to number of cells in each sample; see what kind of coverage we have
** Mapped data/called consensus sequence using BWA and GATK
** Create pileup files from .fixed.bam files, restricted to CCDS regions
* Counted statistics

Revision as of 22:41, 23 September 2011

TO DO

  • DONE - Design LC Sciences Probes - sent probes to Dr. Zhang, Dinh, and Noi
  • Free space on Genome-Miner, create Shared Folder for HiSeq
    • Still need to reformat Ext6T, ideally before September 26
  • Package Exome Scripts for Thomson Lab
  • Barcoding Padlock Probe libraries - talk to Dinh and Noi
  • Low-input Bisulfite Conversion
  • Paper with Sergio
  • Low-input Exomes (Athanasia) - analyze data?
  • Papers for Doug - ask Dr. Zhang?
  • Review GR Manuscript - by September 28

LC Sciences Probes

  • Turned out I made a mistake in probe design yesterday
    • Some probes were allowed to have arms longer than 40 bp total
  • Redesigned probes for same targets (YX Genes [HORMAD1, ZG16] and Ecker mutation/memory that overlaps with PGP1)
  • Obtained 2199 probes that captured at least one CpG and had fewer than 3 CpGs in probe arms
  • Removed targets with high numbers of probes required (due to C's in arms and large size)
    • Also removed 9 additional probes to make the number to order exactly 4000
  • Excel Worksheet (for removal of probes): File:OutputFile LC 09232011.xlsx
  • Probe Set: File:NM LC Probes Filtered 09232011.txt
    • 1820 probes, with 4000 to order due to degeneracy
    • Captures 14,228 CpG sites
  • Created oligos using script: File:Probe2padlockCpgLibSept11 fixed09232011.txt
  • Final oligos: File:NM LC Probes Filtered 09232011.txt.seq

Low-input Exomes

  • Alice used small numbers of Lymphocytes to perform Exome Capture from Nextera libraries
    • Used suppressor oligonucleotides for Nextera adaptor sequences to hopefully increase specificity of capture
    • Used very few cells in each step
  • Need to generate statistics for each library (D, E, F)
    • Currently doing this blind to number of cells in each sample; see what kind of coverage we have
    • Mapped data/called consensus sequence using BWA and GATK
    • Create pileup files from .fixed.bam files, restricted to CCDS regions
  • Counted statistics