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Soil DNA Isolation Kit is suitable for the isolation of total DNA from 500 mg of fresh or frozen stored soil.The total DNA of various microorganisms in the dissolved soil can be bound to the spin column. Digested proteins and humic acid and other PCR inhibitors are filtered out. The genomic DNA is washed by Buffer WB and eluted by Buffer TE, and then it can be used in a variety of molecular biology experiments.Product Composition
Soil DNA Isolation Kit Cat. No. | 5 preps 4102005 | 50 preps 4102050 |
2 ml Sample Tubes Spin Columns Proteinase K Buffer PD Buffer ST Buffer TE Buffer P Buffer WB (concentrate) Instructions | 5 sets 5 sets 120 µl 6 ml 1.2 ml 2 ml 3 ml 1.5 ml 1 copy | 50 sets 50 sets 1.2 ml 55 ml 12 ml 20 ml 28 ml 10 ml 1 copy |
Product Storage and Expiry Date
1． Proteinase K can be transported at room temperature, please store at -20°C after receiving the product.
2． Other reagents and components are stored at room temperature (0~30 ℃), the performance of the product can be maintained without significant change within two years; if the product is stored at 2~8 ℃, the validity period of the product can be extended to more than two years (the product stored at 2~8℃ should be restored to room temperature before use).
Product Description
This product is suitable for the isolation of total DNA from 500 mg of fresh or frozen stored soil. The total DNA of various microorganisms in the dissolved soil can be bound to the spin column. Digested proteins and humic acid and other PCR inhibitors are filtered out. The genomic DNA is washed by Buffer WB and eluted by Buffer TE, and then it can be used in a variety of molecular biology experiments.
Equipment and Reagents to Be Supplied by User
1． Deionized water, absolute ethanol and isopropanol
2． 1.5 ml microcentrifuge tubes and 2 ml microcentrifuge tubes.
3． Pipettes and tips (pipette tips with filter cartridges are recommended to avoid cross-contamination between samples)
4． Protective equipment such as disposable latex gloves and paper towels
5． Microcentrifuge(s) (with rotor for 1.5 ml and 2 ml microcentrifuge tubes)
6． Water bath and homogenizer or vortexer
Preparation Before Use
1． If the centrifuge has refrigeration function, please set the temperature to 25°C.
2． Set the temperature of the water bath to 70℃, incubate Buffer PD, Buffer ST and Buffer TE at 70°C.
3． Add absolute ethanol to Buffer WB according to the instructions on the label of the reagent bottle, and tick the box on the label to mark "Ethanol added".
Protocol:
1. Weigh less than 500 mg of soil and add to a 2 ml sample tube. Add 1 ml Buffer PD, screw the tube tightly and place it in a homogenizer and process for 30 seconds on the full speed; if there is no homogenizer, vortex vigorously on a vortexer for 5 minutes.
* If freeze-dried soil is used, an additional 100 µl of deionized water should be added to facilitate suspension of the soil.
2. Add 20 µl Proteinase K, close the lid and mix well, incubate at 70°C for 15 minutes. Vortex vigorously for 30 seconds every 5 minutes during incubation.
3. Add 200 µl Buffer ST, vortex vigorously for 30 seconds, and centrifuge at 13,000 rpm for 10 minutes.
4. Pipet the supernatant (about 1 ml) into a clean 2 ml microcentrifuge tube (not provided).
5. Add 800 µl isopropanol. Mix well by gently inverting the tube 4~6 times. Centrifuge at 13000 rpm for 10 minutes.
6. Discard the supernatant, centrifuge at 3000 rpm for 5~10 seconds to collect the residual supernatant to the bottom of the tube. Pipet out the residual supernatant and retain the precipitate at the bottom of the tube.
7. Add 100 µl prewarmed Buffer TE and vortex until all the precipitate is dissolved.
8. Add 500 µl Buffer P and mix well by pipetting 6 - 8 times. Pipet the mixture into a spin column, close the lid and centrifuge at 12000 rpm for 30 seconds.
9. Discard the filtrate in the 2 ml collection tube, place the spin column back into the 2 ml collection tube, add 600 µl Buffer WB to the spin column, close the lid and centrifuge at 12000 rpm for 30 seconds.
* The filtrate does not need to be completely discarded, if you want to avoid the contamination of the centrifuge by the filtrate adhering to the nozzle of the collection tube, you can slap the 2 ml collection tube upside down on a paper towel once.
* Make sure absolute ethanol has been added into Buffer WB.
10. Discard the filtrate in the 2 ml collection tube, place the spin column back into the 2 ml collection tube and centrifuge at 14000 rpm for 1 minute.
* If the top speed could not reach 14000 rpm, centrifuge at full speed for 2 minutes.
* Do not omit this step, otherwise, it may cause problems in downstream applications due to the residual ethanol in the eluate.
11. Discard the 2 ml collection tube, place the spin column in a clean 1.5 ml microcentrifuge tube, add 100~200 µl pre-warmed Buffer TE to the center of the membrane, close the lid and incubate for 1 minute at room temperature, then centrifuge at 12000 rpm for 30 seconds.
* If the centrifuge does not have a leak-proof cap, please change the centrifugation condition to 8000 rpm for 1 minute to prevent the 1.5 ml microcentrifuge tube cover from falling off and damaging the centrifuge.
12. Discard the spin column, the eluted DNA can be used immediately for various molecular biology experiments; or store the DNA at -20°C for later use.
* When used for PCR, the DNA should not exceed 1/10 of the final reaction volume (e.g., if the final reaction volume is 50 µl, no more than 5 µl DNA should be added).
