DNA and RNA extraction methods for fungi - Database & Sql Blog Articles

Fungal DNA and RNA extraction methods

The inevitable research on the function of fungal genes always involves DNA and RNA extraction. Because there are more polysaccharide components in the fungus, the quality and subsequent analysis of the DNA and RNA are affected. Therefore, a high-quality, affordable extraction method is our priority. Here, I will share the extraction method I have been using for your reference.

Extraction of fungal DNA (method 1)

1. Take 0.5g of fungal hyphae and quickly grind it into powder in liquid nitrogen.

2. Add 4mL of extract and mix quickly by shaking

3. Add an equal volume of 4mL: isoamyl alcohol (24:1), vortex for 3 to 5 minutes (here is no crude addition of phenol, you can save costs!)

4.1000rpm, 4°C, 5min

5. The supernatant is re-extracted with isoamyl alcohol (24:1) (10,000 rpm, centrifuged at 4 ° C for 5 min)

6. Take the supernatant, add 2/3 volume of -20 ° C pre-cooled isopropanol or 2.5 volumes of absolute ethanol precipitation, mix and let stand for about 30min

7. Pick up the flocculent precipitate with a capillary rod, rinse it repeatedly with 75% ethanol, rinse once with absolute ethanol, blow dry, and resuspend in 500ulTE.

8. Add 1ul of RNaseA (10mg/mL) and treat at 37°C for 1hr.

9. One extraction with phenol (pH 8.0): isoamyl alcohol (25:24:1) and isoamyl alcohol (24:1) (10,000 rpm, centrifugation at 4 ° C for 5 min)

10. Take the supernatant, 1/10V3MNaAc, 2.5V volume of absolute ethanol, precipitate at -70 ° C for more than 30 min.

11. The precipitate was rinsed with 75% ethanol, air dried, dissolved in 200 ul TE, and stored at -20 °C.

Fungal DNA and RNA extraction methods far from biological biological reagents