Oral Swab Genomic DNA Extraction Kit

1. Components of the reagent kit

2. Storage

This reagent kit should be stored at room temperature (15-25℃) and in dry conditions, with a shelf life of 12 months. The DNA extraction purification columns can be stored for 1 year in a cool and dry environment. Proteinase K and RNaza A contain preservatives, allowing transportation at room temperature, but for long-term storage, they should be kept at -20℃.

3. Instructions for Using the Reagent Kit

3.1 This reagent kit is intended for molecular biology research and should not be used for disease diagnosis or treatment.

3.2 Some components in the reagent kit contain irritants. Protective measures such as wearing protective clothing and goggles are recommended.

3.3 During the usage of this reagent kit, a high-speed centrifuge, water bath (metal bath), vortex mixer, anhydrous ethanol, sterile deionized water, and EP tubes need to be prepared by the user.

4. Introduction to the Reagent Kit

The Oral Swab Pročišćavanje DNK Kit provides a rapid and efficient method for purifying DNA from oral swabs, widely used for the extraction of epithelial cells such as oral epithelium.

The Oral Swab DNA Rapid Purification Kit can extract total DNA from oral epithelial cells within 30 minuta. The entire purification process does not require toxic reagents such as phenol-chloroform. The extracted DNA can be directly used for PCR, Southern blotting, and other applications.

5. Experimental Principles and Procedures

6. Extraction Process

Before Starting the Experiment:

A. Reagent Buffer B and C may precipitate under low-temperature conditions. We recommend heating at 65℃for 5 minuta. After the precipitate dissolves, it can be used normally.

B. Before use, add the specified amount of anhydrous ethanol to Wash Buffer 1as indicated on the bottle label. Mark a check on the label to indicate the addition of anhydrous ethanol.

C. Elution Buffer is a0.1x TE solutioncontaining minimal amounts of EDTA. If EDTA might affect subsequent experiments, it is advisable to substitute Elution Buffer with sterile deionized water.

1. Sample Handling:
2. Sampling: Take a sterilized cotton swab, insert it into the mouth, rub against the inner cheek back and forth for more than 20 times.
3. Use scissors to cut the head of the cotton swab, place it into a 2 ml centrifuge tube, and add 400μl of Reagent Buffer B.
4. Dodati10μl Proteinase K (10 mg/ml) and 10μl RNase A (10 mg/ml), invert thoroughly, incubate at 65°C for 20 minuta, vortex for 10 seconds during incubation.
5. Dodati 400μl of Reagent Buffer Cto the lysate, vortex thoroughly.
6. Dodati 200μl of pre-chilled absolute ethanol, mix well; precipitation may occur, but it will not affect subsequent experiments.
7. Transfer the obtained liquid into a DNA extraction and purification column (kit) (approximately 650~700μl each time), centrifuge at >8,000 rpm for 1 minute, discard the collected waste liquid, and reinsert the collection tube into the DNA extraction and purification column (kit) for the next step.
8. Place the DNA extraction and purification column (kit) into a collection tube, add 300μl of Wash Buffer 1, centrifuge at >8,000 rpm for 1 minute, discard the waste liquid, and reinsert the DNA extraction and purification column (kit) into the tube for the next step.

(Note: Ensure that absolute ethanol has been added to Wash Buffer 1.)

7. Dodati 500μl of Wash Buffer 1to the DNA extraction and purification column (kit), centrifuge at 14,000 rpm (20,000×g) za 2 minuta, extend the centrifugation time appropriately to dry the membrane further.
8. Place the DNA extraction and purification column (kit) into a new centrifuge tube, open the lid, incubate at 65°C for 2 minuta; this step can be extended appropriately to evaporate ethanol as much as possible, preventing residual ethanol from affecting downstream experiments.
9. Elute 100μl of Elution Bufferonto the membrane, centrifuge at 12,000 rpm for 2 minuta.

(Note: 1. Eluting DNA with 50μl of Elution Buffer can increase DNA concentration but decrease total DNA yield; 2. Eluted DNA in the eluate can be reapplied to the DNA extraction and purification column, centrifuge again at 12,000 rpm for 2 minutes to increase DNA yield.)