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Stephen Russell

Second Trial - Microwave Extraction

2 posts in this topic

For this test, a variety of different species were tested following the same protocol. 1 minute microwave, 30 minutes heat bath, 10 minutes freezer, 5 minute centrifuge. The 10 uL of the supernatant was used for PCR.

Izumitsu et. al. reported a 93.5% success rate for PCR using the flesh of fresh fruitbodies. Dornte reported a 73.5% success rate. Out of my first 24 specimens, I am looking at 65% sucess rate for fresh specimens (13/20) and 50% for dried Amanitas (2/4). 

I think the next step will be to try to see if some of the additional steps that were included in the first trial will help to remove some of the PCR inhibitors from the negative samples.

mirowave-trial-2.jpg

Top Series

Lane 1 - Ricknella fibula - Negative PCR
Lane 2 - Clitocybe sp.
Lane 3 - Stropharia sp.
Lane 4 - Polyporus leptocephalus
Lane 5 - Ampulloclitocybe clavipes
Lane 6 - Psathyrella sp.
Lane 7 - Panus rudis
Lane 8 - Clitocybe sp. 
Lane 9 - Russula sp. - Negative PCR
Lane 10 - Ladder

Lower Series

Lane 1 - Gymnopus sp. - Negative PCR
Lane 2 - Lentinellus micheneri - Negative PCR
Lane 3 - Psathyrella sp.
Lane 4 - Clitocybe sp.
Lane 5 - Fungi sp. - Negative PCR
Lane 6 - Auricularia sp. 
Lane 7 - Rhizomarasmius sp. - Negative PCR
Lane 8 - Kretzschmaria deusta
Lane 9 - Xylaria sp.
Lane 10 - Marasmius rotula - Negative PCR
Lane 11 - Stereum complicatum
Lane 12 -  Dried Amanita 213772 - Negative PCR
Lane 13 - Dried Amanita 213792 - Negative PCR
Lane 14 - Dried Amanita 215424
Lane 15 - Dried Amanita 213834
Lane 16 - Ladder

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The final 8 of trial 2 came back. 6/8. Brings the total to 19/28. Slightly improves to 68%.Trial 2.jpg

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