StarPure Plasmid Extraction Kit (Membrane Column Method)

1. High efficiency and good quality of plasmid extraction;
2. High purity, no RNA residue;

1. The extraction volume is higher than that of competing products on the market

The DH5α strain containing plasmids of different sizes, 5 kb, 3 kb, and 2 kb, was cultured in LB medium until OD ₆₀₀ ≈2.5, and the plasmid extraction kits from different manufacturers were used for extraction, with 1 mL of bacterial solution for each sample.
The final plasmid elution volume is 50 μL. The concentration and yield of plasmid extracted by Qihengxing are higher than those of other manufacturers.

2. The extracted product is of high purity and free of genomic and RNA contamination.

The DH5α strain containing plasmids of different sizes, 5 kb, 3 kb, and 2 kb, was cultured in LB medium until OD ₆₀₀ ≈2.5, and the plasmid extraction kits from different manufacturers were used for extraction, with 1 mL of bacterial solution for each sample.
The plasmid elution volume was 50 μL. The extracted product was subjected to electrophoresis and had no genomic and RNA contamination and high purity.

Product ingredients:

Self-prepared reagents: Isopropanol, 75% ethanol.

Steps:

The following operation process is based on OD ₆₀₀ Take 1~2 mL of E. coli culture solution ≤4 as an example. If OD ₆₀₀ If the concentration is too high or the amount of bacterial solution increases, the volume of reagents can be increased as appropriate, but the ratio of Resuspension Buffer, Lysis Buffer and Extraction Buffer must be kept unchanged to avoid affecting the extraction effect.

1. Take 1-2 mL of bacterial culture medium and place it in a centrifuge tube. Centrifuge it at 12,000 rpm for 2 min or 4,000 rpm for 10 min, and discard the supernatant completely.
2. Remove the centrifuge tube from the centrifuge, add 250 μL Resuspension Buffer (P1) and 5 μL RNase A, and vortex until the precipitate is completely suspended and the solution becomes white and turbid.
3. Add 250 μL Lysis Buffer (P2), invert several times without vortexing, and let stand for 1 min (the fragmentation state can be observed; if the bacterial concentration is too high, the lysis time can be extended to a maximum of 10 min).
4. Add 250 μL Express Buffer (P3), invert the tube several times, and let it stand for 5 min.
5. Centrifuge at 4℃ 12000 rpm for 10 min or 4000 rpm for 20 min, transfer about 550 μL of supernatant to a clean centrifuge tube for later use, and be careful to avoid the small amount of foam that may remain in the supernatant (excessive foam can be removed by centrifugation again). A small amount of impurities remaining during transfer will be removed by washing in the subsequent steps and will not affect the purity of the product.
6. Add 250 μL of isopropanol to the centrifuge tube (containing the supernatant) in step 5, vortex and mix, then transfer the liquid to the DNA adsorption column (the DNA adsorption column is placed in the collection tube) and incubate at room temperature for 10 minutes.
7. Centrifuge at 12,000 rpm for 1 min, discard the waste liquid in the collection tube, and put the adsorption column back into the collection tube.
8. Add 600 μL 75% ethanol, centrifuge at 12,000 rpm for 1 min, and discard the waste liquid in the collection tube.
9. Repeat step 8 once.
10. Place the adsorption column back into the collection tube and centrifuge at 12,000 rpm for 1 minute. Note: The purpose of this step is to remove the residual ethanol in the adsorption column. The residual ethanol will affect the subsequent enzymatic reactions (enzyme digestion, PCR, etc.).
11. Place the adsorption column in a new collection tube, add 50-100 μL Elution Buffer to the middle of the adsorption membrane, leave at room temperature for 2-5 minutes, centrifuge at 12,000 rpm for 2 min, and collect the plasmid solution in a centrifuge tube. Store the plasmid at -20℃.