FastDNA™ SPIN Kit for Feces

DNA isolation / purification Bacteria - Gram positive Clostridium difficile

Experiment
DNA isolation / purification Bacteria - Gram positive Clostridium difficile
Product
FastDNA™ SPIN Kit for Feces from MP Bio
Manufacturer
MP Bio

Protocol tips

Upstream tips
If humic acid contamination is suspected, repeat the protocol
from the beginning with a fresh stool sample and repeat step #1 once, as well as steps #10 & #11 once each in order.

Publication protocol

DNA extraction
Samples were thawed on ice, homogenized, and 250 mg each were used to extract DNA with commercial kits (Table 1) according to the respective manufacturer's instruction with slight modification as follows: as suggested in the protocol of the FastDNA™ SPIN Kit for Feces and FastDNA™ SPIN Kit for Soil (MP Biomedical, Solon, OH, USA) an extended centrifugation for 15 min at 14,000g was performed after homogenization in the FastPrep®‐24 instrument (MP Biomedical) at 6 m/sec for 40 sec. DES (DNase/Pyrogen‐Free Water) was warmed up at 55°C prior to DNA elution from the SPIN™ Filter. No Proteinase K was added to the NukExPure RNA/DNA (Gerbion, Kornwestheim, Germany) protocol. When using the Precellys Soil DNA Kit (PEQLAB, Erlangen, Germany) lysis was performed using the FastPrep®‐24 instrument at 4 m/sec for 20 sec, followed by centrifugation at 17,000g for 5 min. Lysis with the protocols of the PowerLyzer™ PowerSoil®, PowerFecal™, and PowerSoil® DNA Isolation Kit (Mo Bio Laboratories, Carlsbad, CA, USA) was in all three cases performed with FastPrep®‐24 at 6.5 m/sec for 45 sec. In case of DNA extraction from fecal material using the PowerLyzer™ PowerSoil® and PowerSoil® DNA Isolation Kit an additional centrifugation at 10,000g for 2 min was performed to achieve a pellet. The NucleoSpin® Soil (Macherey‐Nagel, Düren, Germany) protocol was evaluated in four variations of two alternative lysis buffers, SL 1 and SL 2, and in combination with or without the Enhancer SX. Lysis was always achieved using a FastPrep®‐24 at 5 m/sec for 30 sec. To obtain clear supernatant after the lysis of fecal material centrifugation was performed twice for 2 min at 11,000g. The InnuPREP Stool DNA Kit (Analytik Jena, Jena, Germany) was used according to the manufacturer's instructions, whereas the innuSPEED Stool DNA Kit and innuSPEED Soil DNA Kit (Analytik Jena) required an additional centrifugation for 2 min at 13,000g after lysis using the FastPrep®‐24 instrument at 6 m/sec for 40 sec. The QIAamp® DNA Stool mini kit (Qiagen, Hilden, Germany) was used according to the manufacturer's instruction.

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Discussion

Discussion

2 years ago

Author: Israel

How can I improve my DNA yield?

The DNA concentration after using this DNA isolation kit is sometimes too low and thus it is not sufficient for my follow-up experiments. How can I improve it?

Discussion

2 years ago

Author: Milena Alexeyeva Russian Federation

Tips on storing DNA templates?

Hello there! I just started doing experiments on bacterial DNA and I would like your opinion on storing DNA templates. Which are the desired and most optimal conditions?

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing DNA isolation / purification Bacteria - Gram positive Clostridium difficile using FastDNA™ SPIN Kit for Feces from MP Bio.

Paper title
Evaluation of extraction kits and phylogenetic diversity of the porcine gastrointestinal tract based on Illumina sequencing of two hypervariable regions
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Manufacturer protocol

Download the product protocol from MP Bio for FastDNA™ SPIN Kit for Feces below.

Download PDF Download manufacturer protocol

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