The outer layer of fecal bulk was peeled carefully with a sterile scalpel and polystyrene tweezers (approximately 1–2 mm was removed). The inner part of the fecal bulk was used for extraction to avoid a possible contamination with soil organisms and/or the risk of egg deposition by some flies, which would result in the amplification of non-prey organisms. DNA was extracted using a modified version of the Qiagen fecal procedure (QIAamp DNA Tissue Kit, Qiagen Inc., Germany)60 (link). A 200-mg aliquot of each fecal sample was placed in a 2-ml tube containing 200 mg of a mixture of 0.1-, 0.5-, and 2-mm zirconium beads and 1.5 ml of ASL buffer (Qiagen). The sample was bead-beaten at 3200 rpm for 90 seconds, followed by heating at 95°C for 10 minutes. The final pellet was suspended in 180 μl of tissue lysis buffer and incubated with proteinase K for 2 hours at 55°C. The manufacturer's recommendations were followed for the purification and elution of DNA.
Zirconium beads
Manufactured by Qiagen
Zirconium beads are a type of laboratory equipment used for sample preparation and processing. They are small, dense, and chemically inert spherical particles made from zirconium oxide. Zirconium beads are commonly used in various applications, such as mechanical disruption of cells, homogenization of tissues, and bead-beating for DNA/RNA extraction and purification.
Automatically generated - may contain errors
2 protocols using zirconium beads
1
Fecal DNA Extraction Protocol
2
Fecal DNA Extraction from HIV Patients
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Total DNA was extracted from the 13 frozen fecal samples of HIV-infected patients using a modification of the Qiagen stool procedure and the QIAamp® DNA Stool Mini Kit (Qiagen, Courtaboeuf, France) [5 (link)]. Briefly, aliquots of 200 mg of feces were added to tubes containing a 200 mg mixture of 0.1, 0.5, and 2 mm zirconium beads and 1.5 ml of ASL buffer (Qiagen). The sample was bead-beaten at 3200 rpm for 90 seconds, followed by heating at 95°C for 10 minutes. The final pellet was suspended in 180 μl of tissue lysis buffer and incubated with proteinase K for 2 hours at 55°C. Then, DNA was prepared from the solution by using QIAamp spin columns (Qiagen) in an Eppendorf microcentrifuge, following the manufacturer’s instructions.
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