CTR:Notebook/Migrant/2016/04/08

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Whole Genome Sequencing PCR-Free Library Prep for AMKE

  • Used TruSeq Library Prep Kit
  • Diluted sample 1783-83757 to 1ug of DNA in 100 uL total of Buffer AE
  • Sonication (2016-04-06)
    • BioRuptor NGS: 4 cycles of 30 seconds on, 90 seconds off, High Power

1) 2uL 100bp low scale ladder,

3) 2uL sheared AMKE →Sheared DNA to ~400bp.

  • Dried down samples to get concentrated volume of 50 uL using SpeedVac (2016-04-06)
  • Clean-up fragmented DNA (2016-04-07)
  • End Repair and Size Selection- edited to size select for 400bp (2016-04-07)
    • Made a diluted bead mixture with the ratio of 105ul beads: 79ul dH2O and added 160 ul of mixture to each tube

1) 2uL 100bp low scale ladder,

3) 2uL AMKE

  • Adenylate 3' Ends (2016-04-08)
    • Ran on JOHN thermocycler under program: SDCRKAD, Block B
  • Ligate Adapters (2016-04-08)
    • Assigned Adapter 5
  • Gel of Final Library:

1) 2uL 100bp low scale ladder,

3) 2uL AMKE

  • Picogreen:7.32 ng/μL
  • Run sample on BioAnalyzer DNA High Sensitivity Chip (2016-04-18)

  • Sent 10nM to Berkeley for sequencing (Illumina HiSeq 250 PER (Rapid Mode)) ()



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