LA3- Genomics Flashcards
(52 cards)
What did they use first for single genome analysis
Isolates cultured
Then panfenome sequencing of strains eg e0157 stx ecoli
What is metagenomics
Random sequencing of whole popn with a mix of species in a habitat eg the gut
What does this bypass need for
Isolation which has culture bias
What are the steps of wgs of communities (meta genome)
Fragmentation of dna extracted
Computer algorithms assemble it into long reads called scaffolds
Sequence then annotated using information from databases like KEGG or CAZy for gene prediction
What would you use to find the microbes present
16s rRNA amplicon
Amplified with PCR and sequenced
Bioinformatics used to align sequences , and do phylogenetic analysis
What regions of the 16s RRNA will differ between species so can be determined via databases
Hyper variable regions
What is OTU
Organisms with 97% homology
For the question whag are they doing? What would you do
Shot gun wgs
What was used after Robert coch cultivation late 1800s
Amplicon
What was one of the earliest sequencing techniques used for amplicon and what was the advantage
Sanger sequencing
(Dideoxy)
Produced long reads meaning you can get species and genus level due to more hyper variable info
Give another with long reads that then replaced Sanger because it was too expensive
Roche purosequencing
Why did illumina sequencing replace these and what would be an issue theoretically but isn’t because of it being high throughput
Low cost and larger sample depth (could run more at once)
Shorter reads as it looks at 1 hyper variable region only
What does info from HMP do
Produce a reference catalog of genomes from cultures
After illumina hiseq used for metagenomics, what tool is used to run the sequence against databases to identify protein coding genes
Blast search
Give 2 examples of functional databases and what they do
KEGG and cazy
KEGG quantifies abundance of protein genes in a particular pathway eg carb metabolism
Cazy is a metabolic database with info on all enzymes
How are these databases still biased
Biased to functional genes and cultured genomes
What 3 types of study designs are used
Cross sectional - all popn at one time
Longitudinal - repeated observations/samples
Case-control
What allows infants to be colonised
Down reg of tlr
Fr- why would preweaning bifidobacterium be most common
Use lacto-n-neotetraose HMO whcih can’t be used by others eg btheta
Give an example of how baxteroidetes and frimicutes can fluctuate in adulthood
Obesity ob/ob mouse shown firmicutes increased in ratio
Looking at databases like KEGG what other genes were enriched
ABC transporters (firmicutes use), butanoate pathway proteins found abundant in KEGG database on the Ob ob mouse
Galactose metabolism etc
Cazy active firmicutes allowing utilisation of complex glycans = increased energy harvesting
Which archaea was enriched in taxonomy aswell as firmicutes shown by pyrosequencing and why is this relevant
Methanobrevibacter smithii
It is a methanogrn, consumes h2 for better fermention
What do all these enrhciehd pathways and Microbiota suggest in obese models
Better at harvesting energy = energy imbalance
What’s the difference between microbial eukaryote and bacterial abundance in mega genome analysis
Fluctuation/heterogeneity in eukaryotes eg some will have blastocystis presence and some won’t
The bacterial ratio between B and F is consistent usually
