Polysaccharides and Polyphenols Plant Genomic DNA Extraction Kit, 100 T (for Nucleic Acid Extraction)

Product Information

Description/Introduction

The kit is specially designed for the safe, rapid and efficient extraction of high purity genomic DNA from fungal samples and various complex plant tissues that are rich in polysaccharides, polyphenols. The lysis buffer system has been carefully optimized to not only grind and lyse plant tissue samples directly, but also effectively remove impurities such as polysaccharides, polyphenols, and proteins. The entire process can be completed within 1 hour, and the high purity genomic DNA extracted can be used for various molecular biology experiments, including PCR, enzyme digestion reaction, Southern hybridization, RAPD, AFLP, RFLP and others.

Storage and Shipping Conditions

RNase A is shipped with wet ice and stored at -20°C; Other reagents are shipped and stored at room temperature; valid for 12 months.

Product Contents

Before starting (please read carefully)

1. The cryopreserved plant samples should avoid repeated freezing and thawing, otherwise the quality and yield of extracted DNA will be reduced.

2. If Buffer PGLA precipitates, please heat it at 65°C and use it when it is restored to room temperature.

3. Before use, please add 28 mL isopropanol to Buffer GB and mix well.

4. Before use, please add 18 mL absolute ethanol to Buffer GD and 56 mL anhydrous ethanol to Buffer PW.

5. The initial amount of plant tissue samples has a great influence on the extraction effect of gDNA. If the initial amount is too much, the extraction quality and relative yield of gDNA will be reduced. Before starting the experiment, please refer to the following Table1 for usage of Buffer PGLA and Buffer PGLB corresponding to different initial amount of plant tissue samples.

Table 1 Usage of Buffer PGLA and Buffer PGLB corresponding to different initial amount of plant tissue samples

Assay Protocol / Procedures

1. Lysis of plant tissues:

a. (Recommended) Add appropriate amount of Buffer PGLA (recommended usage in Table 1) to a 2.0 mL Nuclease-Free grinding tube (recommended HT-200-M) in advance, and then add 3~4 4 mm stainless steel beads (recommended G0104-200G). Next, quickly transfer 50~100 mg fresh or cryopreserved plant tissue to the grinding tube. Place the grinding tube on the grinder (recommended KZ-5F-3D) and follow the recommended grinding procedure: set the frequency to 70 HZ, grind for 30 seconds each time, repeating the process 15~20 times with a 5 second interval between each grind. If the sample is difficult to grind, such as seeds, you can increase the grinding times to 30 or more. If the tissue is not thoroughly homogenized, it will affect the yield and quality of DNA. Once the grinding is complete, add 10 μL RNase A to the grinding tube, mix the contents upside down and incubate at 65°C for 15 min, mixing upside down every 5 min.

b. Quickly transfer fresh or cryopreserved plant tissue to a 2.0 mL Nuclease-Free grinding tube (recommended HT-200-M) containing 3~4 3 mm zirconia beads (recommended G0203-150G) and pre-cooled with liquid nitrogen. Place the grinding tube on the grinder (recommended KZ-5F-3D) (quickly precool the adapter in liquid nitrogen before placing the grinding tube) and grind until it is completely powdered (if the tissue is not completely ground to powder, it will affect the yield and quality of DNA). Then add appropriate amount of Buffer PGLA (recommended usage in Table1) and put it on the grinder again for 1 min. Once the grinding is complete, add 10 μL RNase A to the grinding tube, mix the contents upside down and incubate at 65°C for 15 min, mixing upside down every 5 min.

2. Add appropriate amount of Buffer PGLB (recommended usage in Table1) to the centrifuge tube, quickly reverse and mix well, and place on ice for 5 min.

3. Centrifuge at 12,000 rpm for 5 min at 4°C, transfer the supernatant to a new 1.5 mL centrifuge tube(if there is still floating matter in the obtained supernatant, it is necessary to centrifuge at 12,000 rpm for 2 min at 4°C again and transfer the supernatant to another new 1.5 mL centrifuge tube).

4. Add Buffer GB of the same volume as the supernatant to the centrifuge tube and mix it upside down.

5. Place DNA Spin Column on Collection Tube, and transfer the mixture to DNA Spin Column. Each addition does not exceed 600 μL, if exceeds, it can be added in batches.

6. Centrifuge at 12,000 rpm for 1 min at room temperature, and discard the filtrate. Put the DNA Spin Column back into the Collection Tube.

7. Add 500 μL Buffer GD to DNA Spin Column, and centrifuge at 12,000 rpm for 1 min at room temperature, then discard the filtrate.

8. Add 600 μL Buffer PW to DNA Spin Column (please add Buffer PW along the pipe wall to help flush the residual salt on the pipe wall), and centrifuge at 12,000 rpm for 1 min at room temperature, then discard the filtrate.

9. Repeat step 8.

10. Put DNA Spin Column into Collection Tube, centrifuge at 12,000 rpm for 2 min at room temperature to remove the residual liquid.

11. Transfer Spin Column to a new Nuclease-free 1.5 mL centrifuge tube, stand at room temperature for 3~5 min, so that the residual ethanol of Spin Column can be evaporate completely.

12. Add 50~100 μL Buffer TE or Nuclease-free Water to the centrifugal tube, stand at room temperature for 5 min (heating Buffer TE or Nuclease-free Water to 65°C is beneficial to improve elution efficiency).

13. Centrifuge at 12,000 rpm for 2 min at room temperature to collect DNA. To get a higher concentration of DNA, you can also add the first eluent back to DNA Spin Column, then stand at room temperature for 5 min and centrifuge for 2 min at room temperature to collect DNA again.

Note

1. Try to use fresh plant tissue to ensure the yield and integrity of genomic DNA.

2. For long-term preservation, the extracted DNA should be eluted with Buffer TE and stored at -80°C.

3. The obtained genomic DNA should avoid repeated freezing and thawing.

4. If the lysis buffer is very viscous, it is recommended to reduce the initial sample size or increase the amount of lysis buffer.

5. For the samples with more water content, the initial sample size can be increased appropriately.

Schedule

The genomic DNA yield extracted from a variety of plant and fungus sample by this kit is shown in the table below. Genomic DNA yield is related to plant species, organs and growth status. The recommended dosage of samples is shown in the table below.

For Research Use Only!

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