Why the 16s does not allow absolute quantification of microbial in microbiology?
I have recently been given a microbial sequencing dataset based on the 16s method, but I have found that this method does not seem to allow for accurate absolute quantification of microorganisms, what factors affect this? Is it some step in the PCR process? Or is it the high throughput sequencing process? And while exploring this question I learnt about qPCR technology, if we have the right markers, is it possible to get the total amount of microbial in a sample and then get the absolute abundance of the community through relative abundance.
The paper below should help answer your questions. With 16S, you have to take into account potential multiple copies of the 16S gene in that organism.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8467167/
Incomplete Data in 16S rRNA Gene Sequencing
The 16S rRNA gene is commonly used for profiling microbial communities because it contains both conserved and variable regions, which allow for the identification and classification of different bacteria. However, 16S rRNA gene sequencing often does not allow for accurate absolute quantification of microorganisms due to several key limitations:
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