19 March 2014

gel extraction

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ddRADseq size selection

As we don’t have access to a Pippin-Prep, performed size selection old school by gel excision.

  • Large gel
    • 150 ul TBE
    • 2.25 g agarose
    • 7.5 ul ethidium bromide
  • Loaded each sample with ladder on both sides
    • ladder
    • JSG001
    • ladder
    • JSG002
    • ladder
  • Ran gel for about 2.5 hours at 120V
  • Excised three gel pieces for each sample
    • 200bp: 275-300bp fragment including 76bp adapter sequence
    • 300bp: 375-400bp fragment including 76bp adapter sequence
    • 400bp: 475-500bp fragment including 76bp adapter sequence
  • Extracted DNA using Qiagen QIAquick PCR Purification
    • surprised to see a lot of DNA in some tubes. enough to make a clear mass that partially blocked column
    • eluted in 10ul H2O and put in freezer
Image of gel

Note slight smear of digested DNA within sample lanes - confirms that there was DNA to excise.

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