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Cascade cell lyses and DNA extraction for identification of genes and microorganisms in kefir grains

Published online by Cambridge University Press:  13 October 2011

Magdalena Kowalczyk ,
Piotr Kolakowski ,
Joanna M Radziwill-Bienkowska ,
Agnieszka Szmytkowska  and
Jacek Bardowski
Magdalena Kowalczyk*
Affiliation:
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland
Piotr Kolakowski
Affiliation:
Danisco Biolacta, Innovation, Olsztyn, Poland
Joanna M Radziwill-Bienkowska
Affiliation:
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland
Agnieszka Szmytkowska
Affiliation:
Department of Pharmaceutical Microbiology, Medical University of Warsaw, Warsaw, Poland
Jacek Bardowski
Affiliation:
Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Warsaw, Poland
*
*For correspondence; e-mail: [email protected]
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Abstract

Kefir is a dairy product popular in many countries in Central Europe, especially in Poland and other countries of Eastern and Northern Europe. This type of fermented milk is produced by a complex population of symbiotic bacteria and yeasts. In this work, conditions for DNA extraction, involving disruption of kefir grains and a cascade of cell lysis treatments, were established. Extraction procedure of total microbial DNA was carried out directly from fresh kefir grains. Using different lysis stringency conditions, five DNA pools were obtained. Genetic diversity of DNA pools were validated by RAPD analysis, which showed differences in patterns of amplified DNA fragments, indicating diverse microbial composition of all the analysed samples. These DNA pools were used for construction of genomic DNA libraries for sequencing. As much as 50% of the analysed nucleotide sequences showed homology to sequences from bacteria belonging to the Lactobacillus genus. Several sequences were similar to sequences from bacteria representing Lactococcus, Oenococcus, Pediococcus, Streptococcus and Leuconostoc species. Among homologues of yeast proteins were those from Candida albicans, Candida glabrata, Kluyveromyces lactis and Saccharomyces cerevisiae. In addition, several sequences were found to be homologous to sequences from bacteriophages.

Keywords

Complex ecosystemskefir grainscascade cell lysisDNA isolationbiodiversity
Type
Research Article
Information
Journal of Dairy Research , Volume 79 , Issue 1 , February 2012 , pp. 26 - 32
Copyright
Copyright © Proprietors of Journal of Dairy Research 2011

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