1. Applicable samples: DNA can be recovered and purified from agarose gels, PCR reaction mixtures, restriction enzyme digestion solutions, plasmid extraction products, kinase reaction mixtures, and other similar systems.

2. Fragment recovery range: Efficiently recovers DNA fragments ranging from 100 bp to 23 kb in length.

3. Recovery rates: For fragments ranging from 100 bp to 10 kb, the recovery rate is 70–90%; for large fragments between 11 kb and 23 kb, the recovery rate is 50–70%.

4. Product Purity: The recovered DNA exhibits high purity and can be directly used in downstream molecular biology applications, including DNA ligation, sequencing, labeling, PCR, and restriction enzyme digestion.

1. Preparation ratio: For a 50‑reaction setup, add 24 mL of 100% anhydrous ethanol (or 26 mL of 95% ethanol) to 6 mL of DNA wash solution; for a 200‑reaction setup, add 96 mL of 100% anhydrous ethanol (or 104 mL of 95% ethanol) to 24 mL of DNA wash solution.

2. Labeling reminder: After adding ethanol, be sure to label the reagent bottle to prevent duplicate additions or omissions.

3. Storage requirements: After preparation, store at room temperature in a sealed container; tightly screw on the cap after each use to prevent ethanol evaporation and maintain washing performance.

4. Failure Diagnosis: After ethanol evaporates, the washing performance will decline significantly. If salt residues remain or the purity is insufficient following washing, verify the ethanol concentration and replenish it promptly.

Standardizing the procedures for sol preparation, sample loading, and elution is essential to ensuring recovery rates; for large DNA fragments, additional optimization of the elution conditions is required.

1. Gel Excision: When excising the gel, remove as much of the excess blank gel as possible to minimize the volume of solubilization buffer and prevent excessive dilution of the sample.

2. Gel dissolution: Add the universal solubilization solution at a 1:3 ratio, and incubate in a 55°C water bath for 10 minutes until the gel is completely dissolved, gently mixing throughout. Maintain the dissolution temperature below 60°C to prevent DNA denaturation.

3. Sample Loading and Centrifugation: The purification column can accommodate a maximum single‑load volume of 800 μL; if the sample volume exceeds this limit, it must be loaded in multiple aliquots and centrifuged separately.

4. Large‑fragment optimization (>10 kb): Preheat the elution buffer to 60–70°C, dispense it onto the column matrix, let it stand for 2 minutes, and then centrifuge; this significantly improves the recovery rate of large fragments. A second elution step can also be performed to increase yield.

1. Common causes: the tip of the purification column contacts the waste liquid at the bottom of the collection tube, leading to salt contamination; insufficient evaporation of ethanol in the wash buffer, resulting in residual ethanol; incomplete gel dissolution, causing tiny agarose particles to contaminate the eluate.

2. Solutions: When removing the purification column, avoid letting the tip touch the bottom of the tube containing waste liquid; after washing, add an additional step of centrifuging the empty column for 2 minutes to thoroughly remove any residual wash buffer; ensure that the gel is completely dissolved before loading it onto the column.

3. Precautions: Store the washing solution in a sealed container and replenish it with ethanol regularly; perform two washes strictly according to the prescribed procedure to ensure thorough removal of impurities.

The elution buffer can be selected according to downstream experimental requirements, and optimizing the elution conditions can enhance nucleic acid concentration and recovery yield.

1. Elution buffer selection: Ultra-pure water can be used for elution, but the pH must be maintained above 6.0; a pH that is too low will reduce elution efficiency. Alternatively, TE buffer (10 mM Tris–HCl, 1 mM EDTA, pH 8.0) may be employed for elution, which is suitable for long-term DNA storage.

2. Minimum elution volume: The minimum elution volume for Column C No. 1 is ≥10 μL; under standard conditions, using 10–15 μL of elution buffer yields high‑concentration DNA.

3. Techniques for enhancing elution efficiency: Re‑adding the first eluate to the purification column for a second elution can improve DNA recovery; preheating the eluate and extending the incubation time both contribute to higher elution efficiency.

4. Precautions: If downstream assays are sensitive to EDTA (e.g., sequencing or certain restriction enzyme reactions), prioritize elution with EDTA‑free elution buffer or ultrapure water.