Products

Insect gDNA Extraction Kit(SpinColumn)

Packing Specification：

Product Introduction:
The unique combination of Binding Buffer and Proteinase K rapidly lyses cells and inactivates intracellular nucleases. Genomic DNA is then selectively adsorbed onto the silica matrix membrane of the spin column under high chaotropic salt conditions. Through a series of rapid washing-centrifugation steps, Inhibitor Removal Buffer and Washing Buffer remove impurities such as cellular metabolites and proteins. Finally, pure genomic DNA is eluted from the silica matrix membrane using low-salt Elution Buffer.

Product Features:
1.The silica matrix membranes in the spin columns are all imported special adsorption membranes, with minimal variation in adsorption capacity between columns and good reproducibility, overcoming the drawback of unstable membrane quality.
2.No toxic reagents such as phenol are required, nor are steps like ethanol precipitation.
3.Fast and simple: operation for a single sample can usually be completed within 30 minutes.
4.Multiple column washing steps ensure high purity. The typical OD260/OD280 ratio ranges from 1.7 to 1.9, and the DNA length can reach 30kb - 50kb, which can be directly used for PCR, Southern-blot, and various restriction enzyme digestion reactions.

Application Scope:
Suitable for rapid extraction of genomic DNA from cells/tissues of various insects, mollusks, arthropods, and other samples.

Operation Steps: 
Tip: Before first use, add the specified amount of absolute ethanol to Washing Buffer WB, mix thoroughly. After addition, mark the corresponding box to indicate ethanol has been added to avoid repeated additions!

1.Grind fresh or thawed insect tissue samples into fine powder in liquid nitrogen or cut into small pieces (cutting into micro-pieces can improve yield). Take 20-50 mg and transfer it to a 1.5 mL centrifuge tube containing 180 μL of Insect Tissue Lysis Buffer TL, mix by pipetting with a large-bore tip.

2.Add 30 μL of Proteinase K solution (20mg/mL), immediately vortex thoroughly to mix.

3.Incubate the lysate in a 55℃ water bath for 1-3 hours or until the tissue is completely digested, gently vortexing several times during incubation to assist lysis.
Optional step: If RNA contamination is significant, add 20 μL of RNase A (25mg/mL) solution after Step 3, vortex to mix, and incubate at room temperature for 5-10 minutes to remove RNA.

4.Add 200 μL of Binding Buffer CB, immediately vortex thoroughly to mix, and incubate at 70℃ for 10 minutes.

5.After cooling, add 100 μL of isopropanol, immediately vortex thoroughly to mix. Flocculent precipitation may occur at this step.

6.Use a 1 mL tip to aspirate the mixture, transfer it to a Spin Column AC (placed in a collection tube), centrifuge at 13,000 rpm for 60 seconds, and discard the waste liquid in the collection tube.
If insoluble tissue debris clogs the pipette tip, gently wipe the tip on absorbent paper to remove the debris; if only a small amount of mixture is aspirated, discard the tip and debris together to prevent clogging of the spin column.

7.Add 500 μL of Inhibitor Removal Buffer IR, centrifuge at 12,000 rpm for 30 seconds, and discard the waste liquid.

8.Add 600 μL of Washing Buffer WB (check if absolute ethanol has been added!), centrifuge at 12,000 rpm for 30 seconds, and discard the waste liquid.

9.Add another 600 μL of Washing Buffer WB, centrifuge at 12,000 rpm for 30 seconds, and discard the waste liquid.

10.Place the Spin Column AC back into the empty collection tube, centrifuge at 13,000 rpm for 2 minutes to completely remove residual Washing Buffer (residual ethanol may inhibit downstream reactions).

11.Transfer the Spin Column AC to a clean centrifuge tube. Add 100 μL of Elution Buffer TE (pre-warming the elution buffer in a 65-70℃ water bath beforehand improves elution efficiency) to the center of the adsorption membrane. Incubate at room temperature for 3-5 minutes, then centrifuge at 12,000 rpm for 1 minute. Transfer the eluted solution back into the spin column, incubate at room temperature for 2 minutes, and centrifuge at 12,000 rpm for 1 minute.
Larger elution volumes result in higher elution efficiency. If higher DNA concentration is required, the elution volume can be appropriately reduced, but the minimum volume should not be less than 50 μL (excessively small volumes reduce elution efficiency and DNA yield).

12.DNA can be stored at 2-8℃ for short-term use, or at -20℃ for long-term storage.