3. Bacterial Genomic DNA Extraction Kit, 50T $ 128

Product Information

Product Description/Introduction

The Bacterial Genomic DNA Extraction Kit is designed to purify genomic DNA from a variety of bacteria including gram-positive/negative bacteria. This system applies a special lysis buffer in combination with DNA preparation membrane to efficiently purify genomic DNA from bacteria. The protocol provides a simple method to achieve the rapid isolation of highly purified genomic DNA and the entire procedure can be accomplished within 20 minutes after cell lysis. Using the kit 10 – 40 μg of highly purified genomic DNA can be extracted from 1 – 5 mL fresh bacterial culture Genomic DNA prepared by this kit is suitable for a variety of applications, such as PCR, Southern blotting, RAPD, AFLP, RFLP and other molecular biology experiments.

Storage and Shipping Conditions

Ship at room temperature; Store at room temperature and keep dry, valid for 12 months; Proteinase K and Lysozyme are stored at -20℃. Lysozyme avoids freeze-thaw cycle. Once thawed, it should be stored at 4℃for long period.

Product Contents

Before Starting (Please read carefully)

1. If a precipitate has formed in Buffer BGL1, heat it at 65°C until the precipitate has fully dissolved.

2. Before first use, add 18 mL of absolute ethanol to Buffer BPD, add 56 mL of absolute ethanol to Buffer PW and mix well separately.

Assay Protocol / Procedures

1. Add 1-5 mL of fresh overnight bacterial culture into a 1.5 mL centrifuge tube, centrifuge at 12,000 rpm for 1 minute, and discard the supernatant. (if there is more bacterial solution, collect the bacterial precipitate into a centrifuge tube through multiple centrifugations.

Note: For Gram-positive bacteria, lysozyme solution should be added to break the cell wall. The method is as follows: add 130 µL of Lysozyme Buffer, 50 µL of Lysozymen, Mix well and incubate it into water bath at 37℃ for 60 minutes. Mix well it upside down every 10 minutes during incubation.

2. Add 200 μL of Buffer BGL1 to the bacteria precipitate, to completely resuspended by pipetting or vortex.

3. Add 20 μL of Proteinase K and 10 µL of RNase A to mixture, mix well by pipetting and incubate at 56℃ for 20 min, The solution should appear clear, and clarify.

Note: After incubation, the solution should become clear. If not, extend the incubation time to 30 minutes, and mix once every 10 minutes.

4. Add 200 μL of Buffer BGL2 and 200 μL of absolute ethanol to the centrifuge tube, mix thoroughly upside and down. white precipitate may be produced at this step.

5. Place the DNA Spin Columns into the Collection Tubes, transfer all the mixture to the DNA Spin Columns.

6. Centrifuge at 12,000 rpm for 30 seconds, discard the flow-through., and put the DNA Spin Columns back into the collection tube.

7. Add 500 μL Buffer BPD to the DNA Spin Columns, centrifuge at 12,000 rpm for 30 seconds, discard the flow-through and put the DNA Spin Columns back into the collection tube.

8. Add 600 μL Buffer PW to the DNA Spin Columns, centrifuge at 12,000 rpm for 30 seconds, discard the flow-through and put the DNA Spin Columns back into the collection tube.

9. Repeat Step 8.

10. Centrifuge at 12,000 rpm for 2 minutes to remove any residue.

11. Put the DNA Spin Columns into a new centrifuge tube, stand at room temperature for 5-10 minutes.

12. Add 50-100 μL Buffer TE or nuclease-free water to the center of the membrane (Pre-heat the Buffer TE or nuclease-free water at 65℃ can improve elution efficiency).

13. Centrifuge at 12,000 rpm for 2 minutes to collect DNA. If more yield is needed, the flow-through can be re-added into the center of the membrane and let it stand for 5 minutes at room temperature and centrifuge at 12,000 rpm for 2 minutes to elute the DNA.

Note

1. Please read the Product Manual carefully before use.

2. Use fresh experimental material to ensure genomic DNA extracted not to be degraded.

3. Do not use too much bacteria is better for fully lysing, otherwise it may affect the yield and purity of genomic DNA.

4. For your safety and health, please wear safety glasses, gloves, or protective clothing.

For Research Use Only!