Dinh Diep

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Daily Lab Notebook[edit]

Notebook

Tools[edit]

wget --user=USERNAME --password='PASSWORD' -c -nH --cut-dirs=2 -U mozilla -r -np --convert-links --reject="index.html*" -e robots=off http://enhancer.sdsc.edu/bli/brandon/Data/
  • Demultiplexing Mi-Seq runs on genome-miner:
 Make a new directory & go into this directory first. Then type the following commands (change MiSeq output folder and date!):
   ln -s /media/LTS_60T/SeqStore2016/MiSeqAnalysis/160419_M00159_0174_000000000-ANULK/Data Data
   cp /media/LTS_60T/SeqStore2016/MiSeqAnalysis/160419_M00159_0174_000000000-ANULK/SampleSheet.csv .
   cp /media/LTS_60T/SeqStore2016/MiSeqAnalysis/160419_M00159_0174_000000000-ANULK/RunInfo.xml .
   bcl2fastq -o 160419_MiSeq --sample-sheet SampleSheet.csv --barcode-mismatches 0
  • Derek's instructions for MiSeq
 Notes from August 24th 2016: http://genome-tech.ucsd.edu/LabNotes/index.php/Dinh/Dinh_2016/NOTES/2016-8-24

Methylation Sequencing Data Repetiore[edit]

  • All methylation data to be stored on genome-miner
  • Data from a publication will be stored under [FirstAuthorLastName][Year]
  • Data from a project will be stored under the project name
  • WGBS (/media/LTS_33T/WGBS_LTS33)
    • Hg19/Ziller2013 - Ziller et al 2013 ( re-analyzed data )
    • Hg19/Heyn2013 - Heyn et al 2013 ( primary data, re-map )
    • Hg19/Xie2013 - Xie et al 2013 ( primary data, re-map )
    • Hg19/NIHRoadMap
    • Hg19/BlueprintProject
    • Hg19/McGillEMC
    • Mm9/
  • RRBS
  • BSPP

Records of all Zhang Lab Methylation data[edit]

Dinh:Methylation_Data_Records Dinh:MONOD_Data_Records

Most recent meeting notes[edit]

  • [1] Current project meeting notes (Updated May 22, 2015)
  • [2] Current project meeting notes
  • [3] Cancer detection project (Updated April 10, 2014)
  • [4] Senate Exam Meeting (Updated February 28th, 2014)
  • [5] Genome partitioning project (Updated January 16, 2013)
  • [6] 5hmC (CIRM) project (Updated January 3, 2013)

Current Projects and Progress[edit]

Dinh's Projects[edit]

  • MONOD
  June2015_MONOD_Efficiencies File:June2015 MONOD ProbesEfficiencies.txt
  BSPP & SeqCap Mapping statistics: File:150209 SN216 mapping summary statistics.xlsx
  Plasma DNA purification [[7]]
  Test 1 ng RRBS protocol [[8]]
  Need to get results from the test protocol first!:
  Generating RRBS libraries from 1.5 ng plasma DNA [[9]]
  Generating RRBS libraries from 10ng normal plasma DNA [[10]]
  Noi's WGBS and BSPP capture experiment on cfDNA from Illumina [[11]],[[12]]
  • 5hmC Experiments
  Samples Processing and Tracking: Dinh:5hmC_Quantification_Samples_Tracking
  Protocol and notes for Lambda controls: Dinh:5hmC_Lambda_Controls_Protocol
  Protocol and notes for oxBS-seq (true 5mC quantification): Dinh:OxBS_Protocol
      H1 ESC (p48 from Sergio): [[13]]
  Protocol and notes for TAB-seq (true 5hmC quantification): Dinh:TAB_Protocol
      Failed trial 1: H1 ESC (p48 from Sergio): [[14]]
      Trial 2: H1 ESC (p48 from Sergio: [[15]]
  Protocol and notes for MAB-seq (5fC and 5caC quantification): Dinh:MAB_Protocol
  • Probes preparation
  Protocol and notes for preparing probes: Dinh:Probes_Prep
  Protocol and notes for preparing probes: Dinh:Probes_Prep_May2015
  • Stoffel production
  Design
  Plasmid construction - first failed attempt: http://genome-tech.ucsd.edu/LabNotes/index.php/Dinh/Dinh_2014/NOTES/2014-5-12
   - I repeated Sanger sequencing twice but was unable to get the Stoffel gene in the sequences.
   - One big issue was my plasmid prep (no column) have too much genomic DNA background which M13 primers can amplify.
   - For PCR/Sequencing, I need to use column to purify my plasmids.
  Plasmid construction - try again: http://genome-tech.ucsd.edu/LabNotes/index.php/Dinh/Dinh_2014/NOTES/2014-6-23
   - I now use NheI digestion site in the middle of the Stoffel gene to screen for my insert. This is much faster than sequencing.
  • Cancer biomarker
  Analysis of GP1 padlock probes capture
  Analysis of MONOD padlock probes capture
  Analysis of RRBS - Dinh:RRBS_MONOD_Analysis/Mapping
  • Genome segmentation
  WGBS data segmentation using N37, Heyn et al (blood), Xie et al (developmental), and Ziller et al (whole tissues, different individuals).
  Mouse WGBS data segmentation using linear regression model and MOABS
  Salk WGBS tissues data : http://genome-tech.ucsd.edu/LabNotes/index.php/Dinh/Dinh_2015/NOTES/2015-10-12
  • 5hmC capture and sequencing
  HOTSPOTS460K capture experiments optimization [16]
  HOTSPOTS460K experiments summary [17]
  • bisReadMapper
Developments:
 CpG/ChG/Chh position calling bugs fixed.
 New version of code will use Bowtie2. NGS Mapper ROC curves: http://lh3lh3.users.sourceforge.net/alnROC.shtml
 Run notes: dinh:COMPUTATIONAL/bisReadMapper
UMI procedure: dinh:COMPUTATIONAL/umi_procedure
  • DMR330K probes
re-normalization: [18] Specificity measures, [19] Probes efficiency measures, [20] [21] Resubsetting strategy,
9 Technical Replicates: [22] QC values, [23] [24] [25] Simulation of STDEV, 
Random tagging of probes: [26] [27] [28]
Design new probes: [29] HMM to determine UMR, LMR, and HMR
  • Protocol optimizations:
[30] [31] Capture with 30K, 
[32] Capture with 97KA/B
[33] Multiplexing tests
[34] Multivariate capture with DMR220K
[35] N2-adapter 
[36] [37] BSPP [38] Training
[39] [40] Normalization capture
[41] [42] DMR220K + EXP1-3 Supressor oligos
[43] Design Probes for Single-End sequencing
[44] [45] [46] Circularization of probes
  • Genemapster/Meangenemachine Updates
[47] Cuda EC
[48] Sanger methylation validation
[49] Installed upgrades
[50] New CUDA installation
[51] Test SOAP3 -- doesn't work on our GPU :(
  • Bisulfite Patch PCR (w/Michelle)
[52] Design
[53] Consider SNPs

Projects with Collaborators[edit]

  • BluePrint (Christoph Bock, CEMM - Vienna)
 11/12/15 Received reference samples from Christoph - Store them in -80C, Rack id: 5B, Box labels: "BluePrint Reference samples - human gDNA"
  - Sample ids: Hct15p, Hct116p, 2N, 4N, 5T, 5N, 6N, 4T, 3N, 3T, 6T, Hct15, Hct116
  • Twins WGBS (Brinda Rana lab, UCSD)
 11/12/15 Mapping HiSeq High Output run - [[54]]
  • Ovarian cancer (Dana Tsui, Cambridge)
 Samples information Dinh:Projects/OvarianCancer
 Screening tumor samples and buffy coat from Cambridge: [[55]]
  • Myelodysplastic syndromes (Rafeal Bejar, Tiffany Tanaka, Brian Reilly, Moores Cancer Center)
 Samples information Dinh:Projects/MDS
  • N37 individual, multiple tissues (Stanford, Billy Li)
    • BSPP (Noi):[56] 1st batch
[57] Genic methylation and expression levels
  • HAPMAP Study (Zhang lab, on grant proposal)
    • BSPP (Noi): [58]
    • WGBS (Alan): ask Alan
Dealing with WGBS data [59] GM12878
Sequence dependent ASM [60] Binomial test and T-test
  • GA - (Glaucoma and optic atrophy?) (Kang Zhang lab)
    • BSPP (Noi): [61] Noi did the experiment while also training two postdocs from Kang's lab.
[62] mQTL on 30 samples, SOM analysis
[63] PCA and HClust
  • Schizophrenia (UCLA, Tina and Dr. Roel)
    • BSPP (Noi): [64] Noi did the experiment while also training Tina over the summer.
Sample identity - males versus females, [65]
Family background substraction, [66]
SibPair using TTest with family null data, [67]
Overlap SibPair with MPO, [68]
SNP Genetic distance plot, [69]
Bis-SNP = more sensitive SNP calling, [70]
  • African Diversity - 48 Methylomes (UPenn collaboration, Sara/Laura)
    • BSPP (Noi):[71] 1st batch [72] 2nd batch, tried to improve QC but they were all too low compared to previously assayed samples.
[73] Analysis with EIGENSTAT software (population structure)
[74] PCA
[75] mQTL
[76] Cleaning and repeat PCA
[77] Checking for reference genome bias
[78] Standardized CpGs quality filter using UPenn48 as an example.
  • 5fC/5caC DIP-seq (University of Northern Carolina Medical School , Yi/Hao/Li)
    • Barcoding and sequencing by Alan:
[79] Genes Region Enrichment analysis, Biotin-labeled dIP 
[80] Antibody dIP
[81] [82][83] Analysis at repeats
[84] 24 sets BOWTIE mapping
[85] [86] [87] [88] BWA PE mapping and peak calling
[89] RNA-seq - repeats mapping
  • Immunogeneticity and reprogramming (Yang Xu lab)
[91] [92] Memory/mutation calling
[93] Compare WGBS
  • Breast Cancer and Diet (Richard, Moores Cancer Center UCSD)
    • Experiments were done in December 2009, January 2010, and February 2010.
[94] Paired Chi-square test 
[95] [96] Compared resequenced datasets with old
[97] [98] mQTL
  • Methylation and metabolic genes
[99] Calculate DMS
  • Memory/Mutation of iPSCs (Salk - Juan Carlos Lab, Sergio)
    • BSPP (Noi): [100] 2nd batch (MSCs, hESCs) [101] 3rd batch, after differentiation
    • BSPP with N2 protocol (Dinh): [102], [103] 1st batch
[104] Memory/mutation calling
[105] CpG localization 
[106] [107] Localization with ChIP data

Finished/dropped projects[edit]

1. Perform quantification of libraries using PhiX 
2. Perform Cluster Generation and Sequencing, Protocol
3. Run pipeline modules for image analysis, base calling, and sequence alignment.

Slides[edit]

most recent ones at bottom