Nongluk (Noi) Plongthongkum

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Labnote

Sample tracking

Current Projects

Targeted bisulfite sequencing

2014 Projects

BSPP capture of Human DMR 460k set

MONOD's project

Sample record

Samples from Moores Cancer Center

  • 2014-03-04: Received 4 aliquots of plasma, serum, and buffy coat from 3 cases from Cancer Center.
    • The samples were used for DNA isolation comparison using four different kits
      • Experimental design & results: [[1]], [[2]], [[3]]
  • 2014-05-27: Received 10 pancreatic adenocarcinoma specimens (2 serum aliquots/case) from Cancer Center (Sharmeela Kaushal).
    • DNA extraction result: [[4]]
  • 2014-06-05: Received 10 pancreatic adenocarcinoma specimens (3 plasma aliquots/case) and 5 tumor tissues from the same patient from cancer center (Sharmeela Kaushal).
    • These samples were derived from the same patients in 2014-05-27
    • DNA extraction result: [[5]]
    • RRBS library preparation of 100ng input DNA from tumor tissue: [[6]]

Samples from Kang Zhang's lab

  • 2014-05-22: Received 30 plasma samples from Kang Zhang's lab
    • Sample info.: [[7]].
    • They are all from normal samples with no cancer history
    • DNA extraction results: [[8]]

FFPE and Frozen Matched Pair Genomic DNA:Human Tumor Tissue: Colon

  • 2014-05-23: Received two tubes of FFPE and frozen matched pair gDNA from Biochain
    • Sample info.
      • Catalog number: D8235090-FP, lot. B609045
      • Tube1: Matched pair - gDNA - human primary tissue: colon. Conc.: 0.58ug/ul or 580ng/ul, amount: 2ug --> expected volume ~3.44ul
      • Tube2: Match pair - gDNA - human primary tissue: colon. Conc.: 0.076ug/ul or 76ng/ul, amount: 2ug --> expected volumed ~26.31ul
        • (2014-06-01) I added TE buffer to make conc. to 50ng/ul (total volume 40ul) based on original conc. and re-measured conc. with Qubit dsDNA HS assay again (1ul).
Sample Original conc. (ng/ul) Estimate volume (ul) Dilute to 50ng/ul (total volume) Add TE buffer (ul) Qubit conc. (ng/ul)
CTT-Frozen 580.00 3.45 40.00 36.55 52.3
CTT-FFPE 76.00 26.32 40.00 13.68 64.8
- CTT = colon tumor tissue
    • RRBS library preparation of 5ng input DNA: [[9]]
    • RRBS library preparation of 100ng input DNA: [[10]]

BSPP capture

  • GP1V4 or MONOD V1 and GP1V6 capture on 5 cancer samples and 3 blood samples (UCLA, SZ 96 sample set): [[11]]
  • MONOD V2 BSPP capture: [[12]]
  • MONOD V3 BSPP capture: [[13]]
  • Amplified DNA from Illumina captured with GP1V4 using Stoffel Fragment and Illumina's polymerase (no positive control, PC): [[14]]
  • Comparison of cancer DNA samples and amplified DNA from Illumina GP1V4 capture using three different DNA polymerases, Stoffel Fragment, Hemo KlenTaq and Illumina's polymerase: [[15]]

scRRBS

  • Experimental design: [[16]]
  • Trial experiment with STD Illumina adaptors and methylated NEBNext adaptors
  • Experiment on flow-sorted nuclei with methylated adaptor from Illumina TruSeq kit
    • Apr#1 experiment: [[20]]
    • Apr#2 experiment: [[21]]
    • Apr#3 experiment: [[22]] --> continue to sequencing (HL166 run)
      • Data analysis: [[23]]

2013 Projects

  • DNA methylation technology comparison (loci-specific, Blueprint project)
    • First batch of 12k probe set TEST experiment: [[24]], [[25]]
    • Record of samples (32 of mandatory (22) and optional (10) samples arrived June 12, 2013) : [[26]]
    • Normalized probe set TEST experiment: [[27]]
  • Parkinson's disease (PD) data set from Burnham Institute
    • BSPP capture: [[28]]
    • Mapping to hg18 (CpG): [[29]]
    • Mapping to hg19 (CpG and non-CpG):


  • Schizophrenia (UCLA, Roel Ophoff):
    • BSPP capture: [[30]]
    • Raw data in genome-miner (There are 4 batches of sequencing data, some of them failed in read2
Batch1 (HiSeq data from UCLA)
Length: 100bp
/media/SeqStore2/110920_UCLA_RO_BSPP
Note: Index 94 was missing from this batch
Batch2
Length: 101bp
 /media/SeqStore2/111005_HL104/UCLA_Blood_SZ
Lane: 8, PE, PE but success only 1 read (read2 failed)
Note: sample s_8_1_Indx28_fixed.txt.gz sequences have been fixed
Batch 3
Length: 100bp
/media/SeqStore2/111112_HL109/UCLA_Blood_SZ
Lane: 4 – 8, PE but success only 1 read (read2 failed)
Lane 1-3 are Rui’ samples and there were some overlapping index to lane 4-8
Batch 4: HL111
Length: 110bp
Lane 4, PE
http://genome-tech.ucsd.edu/LabNotes/index.php/Noi/NOTES/2012-1-2
/media/SeqStore2/111209_HL111/BSPP
    • Mapping
    • Sample identity check by Dinh (checked after combining three batches of sequencing data):
      • Homozygous SNPs and heterozygous SNPs call for X chromosome to identify males vs. females: [[32]]
      • Genetic distance: [[33]]
    • Mid-parent offspring (mpo) analysis
      • Preliminary result by Dr. Zhang: [[34]]
      • Analysis of combining data (4 batches of sequencing data): [[35]]
    • mQTL analysis: [[36]]
      • Multiple test correction: [[37]]
      • mQTL on 5M impute SNPs [[38]]
      • Repeated analysis on no CpG-SNP methylMatrix: [[39]]
    • ASM analysis:
      • Sequence-dependent ASM identification by TTest: [[40]], [[41]]
      • Plot the distance of candidate CpG SNP ASM and correspondence p-value: [[42]]
      • Multiple test correction: [[43]]
    • Regression analysis: [[44]]
      • PCA analysis with control of GA sample from Kang Zhang's lab: [[45]]
      • Multiple test correction: [[46]]
      • Repeat PCA analysis and classified with LDA: [[47]], [[48]], [[49]], [[50]]
    • Reports of data analysis progress:


  • Geographic Astrophy (GA) (UCSD, Kang Zhang) Note: most of the data analysis was performed by Dr. Zhang and Dinh


  • HAPMAP project (HAPMAP PT01, two families, 1362 and 1464)
    • Note: Sample IDs from HAPMAPPT01 start with NA (DNA product) whereas GM represents cell products
    • BSPP capture: [[55]], [[56]], [[57]], [[58]]
    • Mapping to hg19: [[59]]
    • ASM analysis:
      • Preliminary analysis of sequence-dependent ASM: [[60]]
      • Sequence dependent test by Dinh: [[61]]



  • BSPP capture with probes synthesized by LC Sciences (Nature Methods 2012 paper)
    • BSPP capture (1st experiment designed for SE sequencing-> read both ends at once): [[73]]
    • BSPP capture (2nd experiment, improved to library-free protocol and for PE sequencing): [[74]]
    • Summary of mapping of the data generated by library-free protocol (120426_HL118, lane5): [[75]]


  • ASM analysis of WGBS of NA12878
    • Concatenate fastq files as input for ASM pipeline (this step failed because less or cat command cannot print out all data into the same file: [76]
    • Mapping to separate out the reads based on different chromosomes by Dinh: [[77]] --> output files on genome-miner: /media/Ext12T/DD_Ext12T/HL_WGBS_map
    • ASM output data on genome-miner: /home/nplongth/Noi_scratch/ASM_WGBS.NA12878_2012_03_14




  • ASM and ASCM in human cells project (I work on 330k BSPP capture)



  • WGBS of HAPMAP samples by KAPA protocol (epMotion run)


  • LGH-Project, collaboration with Guanghui, Salk Institute
    • BSPP capture: [[100]], [[101]]
    • Note: This batch of experiment also included 5 samples from Dr. Yang Xu's lab
    • Data analysis
      • LGH-projectB (FA-NSC, 1st priority): Data analysis was performed on both genome-miner & genemapster
Mapping data: /home/nplongth/Noi_scratch/GL.project1-2.20120918/LGH-projectB/hg19.mapping
  • Skin microbiome project, started Jan2013
    • Sample information: [[102]]
    • Library preparation rehearsal: [[103]]
    • 72 Library prep (Library ID: NP-SkinMB_N2_Ind17-88_Feb07.2013). The libraries were light sequenced in HL144 lane1 [[104]], [[105]]
    • Select the top 20 libraires from the 72 libraries for Hiseq run [[106]]. Library ID : CW-SkinMB-20SAG-Mar20.2013
    • Prepare sequencing library of low DNA input (1ng): [[107]], [[108]]. Ligation after using CGI protocol to fragment DNA and A-tailing: [[109]]
  • In Situ sequencing, started Mar2013
    • Matt's Project, I worked with Math on probe prep and in vitro padlock probe capture
    • Probe information: [2013 set]
    • Probe preparation: [[110]]
    • Padlock probe capture: [[111]]
  • Post-CoRE fragmentation library construction, started April2013
    • Jeff's microwell MDA samples: [[112]]
    • Eric's MEF cell samples: [[113]]. Library IDs: EC-MEF-Dev7-1-8-May21-2013 & EC-MEF-Dev7-9-16-May21-2013
    • Eric's PGP1#1 cell samples: [[114]]. Library IDs: EC-PGP1-Dev7-1-12-May23-2013 & EC-PGP1-Dev7-13-24-May23-2013
  • epMotion testing
    • AMPure bead purification optimization: [[115]], [[116]], [[117]], [[118]]
    • Bisulfite conversion/Zymo Lightning MagPrep kit optimization:

Protocols


Library construction (with Dinh)

Lab presentation (Journal club/Progress report)