The yield and quality of cellular and bacterial DNA extracts from human oral rinse samples are variably affected by the cell lysis methodology

Sohrabi, Mohsen, Nair, Raj G., Samaranayake, Lakshman P., Zhang, Li, Zulfiker, Aby Hasanat Md., Ahmetagic, Adnan, Good, David and Wei, Ming Q. (2016) The yield and quality of cellular and bacterial DNA extracts from human oral rinse samples are variably affected by the cell lysis methodology. Journal of Microbiological Methods, 122 64-72. doi:10.1016/j.mimet.2016.01.013


Author Sohrabi, Mohsen
Nair, Raj G.
Samaranayake, Lakshman P.
Zhang, Li
Zulfiker, Aby Hasanat Md.
Ahmetagic, Adnan
Good, David
Wei, Ming Q.
Title The yield and quality of cellular and bacterial DNA extracts from human oral rinse samples are variably affected by the cell lysis methodology
Journal name Journal of Microbiological Methods   Check publisher's open access policy
ISSN 1872-8359
0167-7012
Publication date 2016-03-01
Year available 2016
Sub-type Article (original research)
DOI 10.1016/j.mimet.2016.01.013
Open Access Status Not yet assessed
Volume 122
Start page 64
End page 72
Total pages 9
Place of publication Amsterdam, Netherlands
Publisher Elsevier BV
Collection year 2017
Language eng
Formatted abstract
Recent culture-independent studies have enabled detailed mapping of human microbiome that has not been hitherto achievable by culture-based methods. DNA extraction is a key element of bacterial culture-independent studies that critically impacts on the outcome of the detected microbial profile. Despite the variations in DNA extraction methods described in the literature, no standardized technique is available for the purpose of microbiome profiling. Hence, standardization of DNA extraction methods is urgently needed to yield comparable data from different studies. We examined the effect of eight different cell lysis protocols on the yield and quality of the extracted DNA from oral rinse samples. These samples were exposed to cell lysis techniques based on enzymatic, mechanical, and a combination of enzymatic-mechanical methods. The outcome measures evaluated were total bacterial population, Firmicutes levels and human DNA contamination (in terms of surrogate GAPDH levels). We noted that all three parameters were significantly affected by the method of cell lysis employed. Although the highest yield of gDNA was obtained using lysozyme-achromopeptidase method, the lysozyme-zirconium beads method yielded the peak quantity of total bacterial DNA and Firmicutes with a lower degree of GAPDH contamination compared with the other methods. Taken together our data clearly points to an urgent need for a consensus, standardized DNA extraction technique to evaluate the oral microbiome using oral rinse samples. Further, if Firmicutes levels are the focus of investigation in oral rinse microbiome analyses then the lysozyme-zirconium bead method would be the method of choice in preference to others.
Keyword Bacterial cell lysis
DNA
Firmicutes
GAPDH
Oral rinse sample
Total bacteria
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
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