NGS Bioinformatics Workshop1.5 Tutorial – Genome Annotation

April 5th, 2012IRMACS 10900

Facilitator: Richard BruskiewichAdjunct Professor, MBB

Workflow for Today

Prepare to visualize annotationGet a genomic sequence from GenbankRepeat mask it.

Retrieve a genomic sequence…

Retrieve a (relatively small <100kb, eukaryote) genomic sequence clone from GenbankQuery Nucleotide divisione.g. Arabidopsis BAC

clone (HE601748.1)Select FASTASave.. To File.. As “Fasta” (rename?)

Blast is a low hanging fruit…

Use BLAST to quickly survey for similar sequencesMegablast against nucleotide

e.g. HE601748 is closest to A. thaliana chr. 5?Megablast against reference RNA sequence db

Repeat Masking

Upload the clone file to RepeatMasker on the web and run with appropriate parameters:http://www.repeatmasker.org/cgi-bin/WEBRepeatMasker

Save the results (including the masked sequence) to your computer

ab initio Gene Predictions

Genscan:http://genes.mit.edu/GENSCAN.html

Cut and paste results as text to a fileFgenesh:

www.softberry.com

Blast2GOhttp://www.blast2go.com

Annotation workbench, via Gene Ontology (GO) terms. First, save the predicted peptides (e.g. from fgenesh)

need to fix the FASTA headers to assign proper identifiers (could write a script?)

(Java web) start blast2go workbench Load in peptides Do the analysis… e.g. run blastp, GO, annotation,

Interpro, etc. See www.geneontology.org for details on GO http://www.ebi.ac.uk/interpro/ for interpro info

EMBOSS

European Molecular Biology Open Software Suite (EMBOSS):

http://emboss.sourceforge.net Download and install version of interest (e.g.

Linux, Mac OSX, Windows…)Decide what do to:

http://emboss.sourceforge.net/apps/groups.htmlLet’s try a CpG island plot (cpgplot)

Study Genes by Comparative Genomics

JGI Vista toolkit:http://genome.lbl.gov/vista GenomeVistarVista