Athurva Gore/LabNotes/2011-9-23
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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