Quantifying Microbial Populations in Samples

The PCR primers used for real-time qPCR of total bacteria [34 (link)], fungi [34 (link)], protozoa [35 (link)], methanogens [36 (link)], and SRB [37 (link)] are listed in Table S1. Real-time PCR was performed on a StepOnePlus system (Applied Biosystems, California, USA) using the SYBR Premix Ex Taq dye (Takara Bio Inc.). Copies of 16S rRNA gene (total bacteria), 18S rRNA gene (fungi and protozoa), methyl coenzyme-M reductase alpha subunit gene (mcrA, for methanogens), and dissimilatory sulfite reductase alpha subunit gene (dsrA, for SRB) in each sample was quantified in three technical replicate against respective standards, which were purified PCR products of known length and concentration. The absolute abundance of each microbial population was expressed as copies of the target gene/g of samples.

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Shen J., Li Z., Yu Z, & Zhu W. (2020). Effects of dietary replacement of soybean meal with dried distillers grains with solubles on the microbiota occupying different ecological niches in the rumen of growing Hu lambs. Journal of Animal Science and Biotechnology, 11, 93.

Publication 2020

StepOnePlus system SYBR Premix Ex Taq dye

18s rrna Alpha subunit Bacteria Dissimilatory sulfite reductase Fungi Gene Methanogens Methyl coenzyme m reductase Primers Real time pcr Replicate Rrna gene

Corresponding Organization :

Other organizations : Nanjing Agricultural University, The Ohio State University

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Copies of 16S rRNA gene (total bacteria)
Copies of 18S rRNA gene (fungi and protozoa)
Copies of methyl coenzyme-M reductase alpha subunit gene (mcr A, for methanogens)
Copies of dissimilatory sulfite reductase alpha subunit gene (dsr A, for SRB)

control variables

None explicitly mentioned

controls

Respective standards, which were purified PCR products of known length and concentration, were used for quantification of the target genes in each sample.

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