Invitrogen™ PureLink™ Microbiome DNA Purification Kit

For highly pure samples, both fluorometers (such as the Qubit Flourometric Fluorometer) and spectrophotometers (such as NanoDrop One Spectrophotometer) are successfully utilized and give very similar readings. However, when dealing with DNA or RNA samples that have high levels of contaminants, such as salts or protein, the Qubit Fluorometer provides a more accurate reading, as this instrument uses dyes that are very specific to the analyte of interest. We have seen some challenging microbiome and FFPE samples where the Qubit Fluorometer readings were up to 2 times lower compared to the NanoDrop spectrophotometer, which is overestimating DNA concentrations. More and more laboratories are use both instruments side by side, as they provide complimentary information: the Qubit fluorometer is more accurate and specific in terms of determining nucleic acid concentration, while the NanoDrop spectrophotometer provides a valuable information regarding the purity (ie., A260/280, A260/230).

We tested 3 liquid versions of transport media: Amies, Stuart, and Cary Blair, with all three being compatible with the PureLink Microbiome DNA Purification kit.

Yes, the PureLink Microbiome DNA Purification Kit works for swabs samples. However this kit does not come with swabs. You will find some suggested swab part numbers that we have tested with this kit in swab protocol on the product page. (http://www.thermofisher.com/order/catalog/product/A29790).

The PureLink Microbiome DNA Purification Kit was developed to be used with a wide range of sample types including stool, soil, urine, saliva, swabs, and other samples. In addition to testing bacterial samples, we have also tested fungi samples. Specialized beads in the kit efficiently break open cell walls and membranes with the help of heat and chemical lysis. This triple lysis approach works great for tough-to-lyse species such as fungi. Additionally, the stool-derived inhibitors are rapidly eliminated with a novel clean up buffer, resulting in DNA that is compatible with any type of downstream analysis.

Yes, when using microbial DNA protocol with PureLink Microbiome DNA Purification Kit, you will get both host and microbial DNA. The bulk of DNA in human stool is of microbial origin. Depending on the sample, we roughly estimate that approximately 10-20% of the total DNA isolated is host DNA while the remaining 80-90% is microbial DNA.

We simplified the protocol for those scientists who are only interested in human DNA isolation from stool, as mammalian cells are easy to lyse and DNA isolation can be performed faster. Microbes have thicker cell membranes of a different composition, and are generally more difficult to lyse. Thus, separate protocols, with additional steps to ensure efficient disruption for isolation of microbial DNA, were developed.

If fresh stool, body fluids, swabs, and other samples cannot be processed, freezing the samples at -20 degrees C or -80 degrees C is the best way to preserve them. We recommend keeping freeze/thaw cycles to a minimum. For many samples, storage at 4 degrees C for up to one week is also fine. Please note, use of commercially available DNA stabilizing reagents can negatively impact the downstream nucleic acid isolation, or make the workflow more challenging or lengthy.

Yes, the PureLink Microbiome DNA Purification Kit has been successfully used for isolation of microbial DNA from human, mouse, rat, horse, cow, dog, cat, and other types of stool/feces. Despite the fact that these samples are very different in terms of consistency, presence of miscellaneous food debris, composition of the microbial community, various inhibitors present, and other variables, the kit enabled fast and efficient isolation of substantial amounts of highly clean microbial DNA.