Project:Plant species testing: Difference between revisions
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We used two methods of extraction. Method 1: | We used two methods of extraction. Method 1: | ||
* Blend peas with salt and detergent | |||
* Incubate for 15 mins at 60 C | |||
* 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: | Method 2: | ||
- 'Homogenize' with a pestle in 5% chelex solution for 1 min | - 'Homogenize' with a pestle in 5% chelex solution for 1 min | ||
- Vortex for 10s then boil for 5 mins, then vortex again for 10s | - Vortex for 10s then boil for 5 mins, then vortex again for 10s | ||
- Centrifuge for 1 min - recover supernatant and use as template | - Centrifuge for 1 min - recover supernatant and use as template | ||
Revision as of 19:53, 17 July 2012
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
- 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
