19 March 2014

gel extraction

Next post Previous post

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

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