Eukaryotic unicellular organisms ar

Eukaryotic unicellular organisms are an important part of the soil microbial community, but they are often
neglected in soil functional microbial diversity analysis, principally due to the absence of specific investiga-
tion methods in the special soil environment. In this study we used a method based on high-density centri-
fugation to specifically isolate intact algal and yeast cells, with the aim to analyze them with flow cytometry
and sort them for further molecular analysis such as deep sequencing. Recovery efficiency was tested at low
abundance levels that fit those in natural environments (104to 106cells per g soil). Five algae and five yeast
morphospecies isolated from soil were used for the testing. Recovery efficiency was between 1.5 to 43.16%
and 2 to 30.2%, respectively, and was dependent on soil type for three of the algae. Control treatments without
soil showed that the majority of cells were lost due to the method itself (58% and 55.8% respectively). However,
the cell extraction technique did not much compromise cell vitality because a fluorescein di-acetate assay indi-
cated high viability percentages (73.3% and 97.2% of cells, respectively). The low abundant algae and yeast
morphospecies recovered from soil were cytometrically analyzed and sorted. Following, their DNA was isolated
and amplified using specific primers. The developed workflow enables isolation and enrichment of intact auto-
trophic and heterotrophic soil unicellular eukaryotes from natural environments for subsequent application of
deep sequencing technologies.

ผลลัพธ์ (อังกฤษ) 2:

Unicellular eukaryotic organisms are an important soil microbial Part of the Community, but they are often
Neglected in soil microbial functional Diversity Analysis, Due principally to the absence of specific Investiga-
tion methods in the special soil Environment. In this Study we used a method based on High-density Centri-
Fugation to specifically isolate Intact algal cells and yeast, with the AIM to Analyze them with flow cytometry
and sort them for further molecular Analysis such as Deep sequencing. Recovery efficiency was tested at low
levels that Abundance Natural Fit in those environments (104to 106cells per soil G). Five algae and yeast Five
Morphospecies isolated from soil were used for the testing. Recovery efficiency was between 1.5 to 43.16%
to 30.2% and 2, respectively, and was dependent on soil for Type Three of the algae. Control treatments Without
Majority of soil Showed that the cells were Lost Due to the method itself (58% and 55.8% respectively). However,
the extraction Cell Technique did not compromise much because a fluorescein di-acetate Cell Vitality assay Indi-
cated viability High percentages (73.3% and 97.2% of cells, respectively). The low Abundant algae and yeast
Morphospecies recovered from soil Cytometrically were analyzed and sorted. Following, their DNA was isolated
and amplified using specific primers. Developed Workflow enables the isolation and enrichment of Intact auto-
trophic and heterotrophic soil unicellular eukaryotes from Natural environments for Subsequent Application of
Deep sequencing Technologies.