Project:Plant species testing

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Revision as of 20:44, 17 July 2012 by Mycoplasma (talk | contribs)
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We are using primers for plants to test and refine our DNA extraction / PCR technique. First project is to test for peas. Inspiration in this paper.


First extraction and PCR - 17/7/12

We used two methods of extraction. Method 1:

  • Blend peas with salt and detergent
  • Incubate for 15 mins at 60 C (optional step not taken)
  • Filter and centrifuge - recover supernatant
  • Add proteinease K (0.8 ul - incubate for 1 hr at 50C then 94 C for 10 mins (inactivation of proteinase K)
  • Precipitate with rubbing alcohol and meat tenderiser as an enzyme
  • Centrifuge and remove supernatant to leave pellet

Method 2:

- 'Homogenize' with a pestle in 5% chelex solution for 1 min - Vortex for 10s then boil for 5 mins, then vortex again for 10s - Centrifuge for 1 min - recover supernatant and use as template

Using the PCR machine:

For soak: (

For PCR:

  • File 4 enter
  • Step
  • temp
  • time
  • temp a
  • time
  • time e
  • Segment 4 - 0 C for 0 seconds
  • Cycle count
  • Link to stored file - 20 C (to keep