MaXtract High Density (100 x 15 ml)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Extraction of nucleic acids from organic solvents

Protocol tips
Extraction of nucleic acids from organic solvents

Publication protocol

Nucleic acid extraction from mouse small intestines. Distal halves of the small intestine from 4 mice from each group were dissected, flash frozen in liquid nitrogen and stored at -80oC. For extraction, each frozen gut section was transferred with sterile forceps to a 2 mL Lysing Matrix E tube (MP Biomedicals) containing 50 μL of 0.1M aluminum ammonium sulfate. Equal volumes (500 μL) of modified CTAB buffer (10% CTAB, 250 mM phosphate, 300 mM NaCl) and phenol:chloroform:isoamylalcohol (25:24:1) were added to each tube, tubes were agitated with a FastPrep (MP Biomedicals: 30 seconds, 5.5 m/s) and centrifuged (5 min, 4oC, 16,000 x g). Aqueous phases were removed by pipette, transferred to phase-lock gel tubes (MaXtract High Density Gel Tubes, Qiagen) containing approximately 1 aqueous volume of chloroform, inverted by hand and centrifuged again to yield a crude nucleic acid extract. Another 500 μL aliquot of modified CTAB buffer was added to the Lysing Matrix E tube and tubes were agitated and centrifuged again as described above. This second aqueous extract was purified with chloroform as above to yield an additional crude nucleic extract. This re-extraction step was repeated twice. Each of the three crude nucleic acid extracts from a sample was transferred from the phase-lock gel tube to an individual 2 mL tube, gently mixed by pipette with 2 volumes of polyetheleneglycol/salt solution (30% wt/vol PEG 6000, 1.6M NaCl), and incubated at room temperature for 2 hours. Crude nucleic acid extracts were then centrifuged (10 min, 4oC, 16,000 x g), supernatants were removed by pipette and pellets were washed with 1 mL ice-cold 70% ethanol. Pellets were resuspended in 60 μL of nuclease-free (DEPC treated) water. The three crude nucleic acid extracts from each sample were combined and a 60 μL aliquot was purified by column chromatography using an DNA/RNA Allprep Kit (Qiagen, CA). DNA and RNA were separately eluted and purified. Purified DNA was eluted in 2 x 25 μL Buffer EB and used as genomic DNA template for PCR amplification.

Full paper   Login or join for free to view the full paper.

Reviews

Discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing DNA isolation / purification Tissue - small intestine using MaXtract High Density (100 x 15 ml) from Qiagen.

Manufacturer protocol

Download the product protocol from Qiagen for MaXtract High Density (100 x 15 ml) below.

Download manufacturer protocol

Videos

Check out videos that might be relevant for performing DNA isolation / purification Tissue - small intestine using MaXtract High Density (100 x 15 ml) from Qiagen. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.