The Metagenome Flashcards
What is genomics?
study of the whole human genome
What is Transcriptomics?
whole cell gene expression
What is Proteomics?
whole cell protein content
What is metabolomics?
The study of whole cell metabolite content
What is metagenomics?
study of genetic material recovered directly from environmental or biological systems/ compartments
What is the Microbiome?
a microbial community occupying a well defined habitat
What is a microbiota?
ecological community of commensal and pathogenic microorganisms
How many variable regions is the 16S ribosome divided into?
9
What is the method for 16s targeted PCR amplification
Sample collection, DNA extraction, 16s PCR amplification, sequencing and analysis
What do you choose the variable region based on?
the phylogenetic signal and Amplicon length
What does your choice of region determine?
Resolution
How do you mitigate contamination?
Randomising samples, Note batch numbers of reagents and sequencing negative controls
Why is targeted 16S PCR amplification biased?
Because it only affects bacteria
What is targeted 16S PCR amplification used for?
To assess taxonomic diversity in the sample
Is whole genome shotgun sequencing biased?
No
How do you analyse samples in whole genome shotgun sequencing?
Either build an assembly of the sequences or bin them (sort them into groups based on similarity)
What is whole genome shotgun sequencing used to do?
Assess composite gene functions in the sample
What are some issues with whole genome shotgun sequencing?
Host cells often in excess in the sample, no amplification step to enrich for bacterial DNA and Sample dependant
How can you enrich without amplification?
Pre or post extraction
How do you enrich the sample before whole genome shotgun sequencing?
differential lysis of Mammalian Cells, Enrich for intact microbial cells
What does pre-extraction enriching DNA in whole genome shotgun workflow cause?
potential bias towards gram-positive bacteria
How do you enrich the sample after whole genome shotgun sequencing?
Enzymatic degredation of methylated nucleotides that targets mammalian DNA
What does pre-extraction enriching DNA in whole genome shotgun workflow cause?
Bias against AT rich bacterial genomes
What are the four metagenomic applications?
Environmental, animal, clinical diagnostics and public health
What is metagenomics used to find out in the environment?
The diversity and relative abundance of organisms
What does MALDI stand for?
Matrix assisted laser desorption/ionisation
Why can’t we always use MALDI?
Lots of things cant be cultured
What is MALDI used to do?
Identifying hard to culture organisms in patient samples and antibiotic resistant repertoires from clinical samples
How are metagenomics used in public health?
Infection control and outbreak management and surveillance of antimicrobial resistance in the food supply
