RIAM: A Universal Accessible Protocol for the Isolation of High Purity DNA from Various Soils and Other Humic Substances

A single universal open protocol RIAM (named after Research Institute for Agricultural Microbiology) for the isolation of high purity DNA from different types of soils and other substrates (high and low in humic, clay content, organic fertilizer, etc.) is proposed. The main features of the RIAM prot...

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Bibliographic Details
Published inMethods and protocols Vol. 5; no. 6; p. 99
Main Authors Pinaev, Alexander G, Kichko, Arina A, Aksenova, Tatiana S, Safronova, Vera I, Kozhenkova, Elena V, Andronov, Evgeny E
Format Journal Article
LanguageEnglish
Published Switzerland MDPI AG 16.12.2022
MDPI
Subjects
Acids
Agricultural chemicals
Alcohol
Biotechnology industry
Clay
CTAB DNA precipitation
Deoxyribonucleic acid
DNA
Ethylenediaminetetraacetic acid
Genetic testing
high phosphate
Humic acid
Minerals
Molecular weight
Phosphates
Polyethylene glycol
Polymerase chain reaction
Protocol
soil DNA isolation
Soils
Sorption
United States
United States > US
soil DNA isolation
high phosphate
CTAB DNA precipitation
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Summary:A single universal open protocol RIAM (named after Research Institute for Agricultural Microbiology) for the isolation of high purity DNA from different types of soils and other substrates (high and low in humic, clay content, organic fertilizer, etc.) is proposed. The main features of the RIAM protocol are the absence of the sorption-desorption stage on silica columns, the use of high concentrations of phosphate in buffers, which prevents DNA sorption on minerals, and DNA precipitation using CTAB. The performance of RIAM was compared with a reference commercial kit and showed very good results in relation to the purity and quantity of DNA, as well as the absence of inhibitory activity on PCR. In all cases, the RIAM ensured the isolation of DNA in quantities much greater than the commercial kit without the effect of PCR inhibition up to 50 ng DNA per reaction in a volume of 15 µL. The latter circumstance along with the ability of the protocol to extract low molecular weight DNA fractions makes the method especially suitable for those cases where quantitative assessments, detection of minor components of soil microbiota, and completeness of isolation of all DNA fractions are required.
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ISSN:2409-9279
2409-9279
DOI:10.3390/mps5060099