User:R. Eric Collins/GenomicsTutorial/Genomics

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7 February 2011 -- Genomics Tutorial -- McGill University

Genome Sequencing

Instrument: Roche 454

Each full 454 plate generates about 300-500 Mbases of raw sequence data

454 Video

Usage Scenario: hypervariable tag sequencing (e.g. V6 of bacterial 16S rRNA) on environmental samples
  • add Titanium fusion primers to your PCR primers
  • barcode many (>48) samples for sequencing as a pool in the same plate
  • send off the pool of samples
  • total cost is $7,800.00
Usage Scenario: de novo genome sequencing of 6 bacterial isolates (5Mb) at 10--12x coverage
  • the most cost effective way to perform this experiment is to barcode these samples and sequence them as a pool.
  • total costs per set of 6 samples is $12,000.00, including library preparation and sequencing.
de novo Assembly
Assembly with Reference Genome
  • whole genome alignment: MUMmer

View reads against reference

Sequence Clustering

Algorithms
Orthologous Groups
Metabolic Pathways/Predictions


Phylogenetics

Find a gene of interest
  1. KEGG or IMG or NCBI
  1. Find the paper at NCBI: An ice-binding protein from an Antarctic sea ice bacterium
  2. All Links From This Record --> Protein
  3. Analyze this sequence --> Run BLAST
  4. Run against 'nr' database using default parameters
  5. click [Distance tree of results]
Get an alignment
  1. IMG or NCBI or EBI
Infer a Phylogeny
  1. IMG or EBI
View a Phylogenetic Tree
  1. Jalview or iTol
Multiple Sequence Alignment
Phylogenetic Inference
Distance Methods
Maximum Parsimony
Maximum Likelihood
Bayesian Inference
Phylogenetic Tree Visualization
Usage Scenario: Bacteria 16S phylogeny

Evolution

Selection