Dual Genomic DNA Isolation Kit (Plant)

Catalog numberPDC05-0100

CategoriesColumn Based

Size100 Reactions

Sample4 rxns ($21.6)

$369.6 Inquire

Note: PureDireX / Purification Kits / Extraction Kit / Plant

Description

The Dual Genomic DNA Isolation Kit (Plant) combines reagent system and spin column system. The kit is designed specifically for isolating genomic DNA from plant samples. This unique reagent system ensures the total DNA with high yield and good quality from samples. The spin column system is designed to purify or concentrate DNA products which have been previously isolated with the reagents. The entire procedure can be completed in 1 hour without phenol extraction. Purified DNA is suitable for use in PCR or other enzymatic reactions.

Sample: 100 mg of fresh plant tissue or 50 mg of dry plant tissue
Format: Reagent and spin column
Operation time: within 1 hour.
Elution volume: 50-200 ul.

Kit Contents

Contents	PDC05-0100	PDC05-0100S (Sample)
Buffer PG	100 ml	4 ml
Buffer BD	100 ml	4 ml
Buffer W1	45 ml	2 ml
Buffer W2 (Add ethanol)	15 ml (60 ml)	300 µl × 2 (1.5 ml × 2)
Buffer E	10 ml	1 ml
Columns DGP	100 pcs	4 pcs
Collection Tubes	100 pcs	4 pcs

Features

Delivering high-quality genomic DNA with the fast procedure.
Ready-to-use genomic DNA for high performance in any downstream application.
Highly purified and high yield genomic DNA can be extracted from various plant samples.
Optimized plant lysis buffer for the efficient lysis.

Application

Gene cloning
Quantitative real time PCR

Southern blotting
SNP genotyping

Protocol

Step 1 Sample Preparation

1. Cut off 100 mg of fresh plant tissue or 50 mg of dry plant tissue.
2. Grind the sample under liquid nitrogen to a fine powder using a mortar and pestle.

Step 2 Lysis

1. Add 1 ml of Buffer PG and 0.5 μl of RNase A (50 mg/ml) to the sample in the mortar and grind the sample until it is completely dissolved.
2. Transfer the dissolved sample to a 1.5 ml microcentrifuge tube.
3. Incubate at 75°C for 30 minutes.(invert the tube every 10 minutes)
4. Centrifuge at 14-16,000 x g for 5 minutes.
5. Transfer carefully the clear supernatant to a new 1.5 ml microcentrifuge tube.

Step 3 Phase Separation

1. Add 600 μl of chloroform to the supernatant from Step 2.
2. Shake vigorously and then centrifuge at 14-16,000 x g for 10 minutes.
3. Carefully remove the upper phase and transfer it to a new 1.5 ml microcentrifuge tube.
4. Repeat the Phase Separation Step until the interphase becomes clear then transfer the clear upper phase to a new 1.5 ml microcentrifuge tube.

*The number of repetitions is dependent on sample type; e.g. dense tissue samples may require a higher number of repeats.

Step 4 DNA Precipitation

1. Add 800 μl of isopropanol to the 1.5 ml microcentrifuge tube containing the clear upper phase from step 3.
2. Mix the sample by inverting gently and let stand for 5 minutes at room temperature (DNA precipitation can be increased with extended standing time).
3. Centrifuge at 14-16,000 x g for 15 minutes.
4. Discard the supernatant and wash the pellet with 1 ml of 70% ethanol.
5. Centrifuge at 14-16,000 x g for 5 minutes.
6. Completely discard the supernatant and re-suspend the pellets in 200 μl of TE Buffer or ddH2O.
7. Incubate for 10 minutes at 75°C to dissolve the pellet.
8. If more pure DNA is required, perform this optional DNA Pure Protocol. View More↘

＊ Add 60 ml of the absolute ethanol to the Buffer W2 prior to initial use.
＊ Pre-heat the Buffer E to 75°C prior to use.

Step 1 Sample Prep.

Add 1 ml of Buffer BD to the sample which have been previously isolated using reagents and shake vigorously.

Step 2 DNA Binding

1. Place a Column DGP in a 2 ml Collection Tube.
2. Transfer the sample mixture from the previous step into the Column DGP.
3. Centrifuge at 14-16,000 x g for 30 seconds.
4. Discard the flow-through and place the Column DGP back in the same Collection Tube.
5. Centrifuge at 14-16,000 x g for 30 seconds.
6. Discard the flow-through and place the Column DGP back in the same Collection Tube.

Step 3 Wash

1. Add 400 µl of Buffer W1 into the Column DGP.
2. Centrifuge at 14-16,000 x g for 30 seconds.
3. Discard the flow-through and place the Column DGP back in the same Collection Tube.
4. Add 600 µl of Buffer W2 (ethanol added) into the Column DGP.
5. Centrifuge at 14-16,000 x g for 30 seconds.
6. Discard the flow-through and place the Column DGP back in the same Collection Tube.
7. Centrifuge again for 3 minutes at 14-16,000 x g to dry the column matrix.

Step 4 DNA Elution

1. Transfer the dried Column DGP to a clean 1.5 ml microcentrifuge tube.
2. Add 50-200 µl of Pre-Heated Buffer E or TE Buffer into the center of the column matrix.
3. Let stand at 75ºC for 5 minutes.
4. Centrifuge for 2 minutes at 14-16,000 x g to elute the purified DNA

Caution

Check buffers before use for salt precipitation. Re-dissolve any precipitate by warming up to 37°C.
During operation, always wear a lab coat, disposable gloves, and protective equipment.
Research Use Only. Not intended for any animal or human therapeutic or diagnostic uses.

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