User:R. Eric Collins/GenomicsTutorial/Genomics

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 Primers: CBDB NCBI
 * Annotation Pipelines: MicroScope MiGAP DIY Genomics IMG/mer JCVI Service BASys GenDB SABIA

Assembly with Reference Genome

 * whole genome alignment: MUMmer

View reads against reference

 * BAMview alignment viewer

Sequence Clustering

 * Sequence Clustering

Algorithms

 * BLAST
 * blast-clust
 * MCL and TribeMCL

Orthologous Groups

 * COG database
 * eggNOG database
 * MEGAN

Metabolic Pathways/Predictions

 * KEGG
 * KEGG Automated Annotation Server
 * MetaCyc metabolic cycles

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

 * List of msa software
 * CLUSTALW @EBI
 * MUSCLE @EBI
 * MAFFT @EBI

Phylogenetic Inference

 * Phylogenetic Inference

Distance Methods

 * CLUSTALW @EBI
 * PHYLIP -- FITCH and NEIGHBOR

Maximum Parsimony

 * fastDNAml
 * PHYLIP -- DNAML and PROML

Maximum Likelihood

 * PHYLIP -- DNAPARS and PROTPARS

Bayesian Inference

 * MrBayes

Phylogenetic Tree Visualization

 * Interactive Tree of Life
 * JalView

Usage Scenario: Bacteria 16S phylogeny

 * Ribosomal Database Project
 * SILVA
 * Greengenes

Selection

 * dN/dS ratio
 * Adaptive Evolution Server