1. Introduction to Bioinformatics - Tutorial no. 13 Probe Design Gene Networks

2. Probe design problem Given a genome with a set of genes. Recall that each gene is represented by a probe (a short segment) of the corresponding cDNA. The length of a probe can be 20, 50, 70! The probe design problem tries to identify a probe for every gene.

3. Probe design problem (Informal definition) Given a genome, for every cDNA (or mRNA) X i,  a probe p i is identified such that p i is a substring of X i ; and p i does not cross-hybridize with other mRNAs Probe design problem creates a probe p i for every mRNA X i X2X2 X3X3 X4X4 X1X1 p1p1 p1p1 p1p1 p1p1 XXX

4. Primer3 Input Sequence 5’ PCR primer Oligo probe 3’ PCR primer Include in PCR primer Exclude from PCR primer Repeats library

5. Primer3 Output mRNA position Length Hybrid T m % GCSelf complementarity 3’ complementarity Sequence Product 5’ PCR primer Oligo probe 3’ PCR primer

6. OligoDB Sequence name to find

7. OligoDB Input Oligonucleotide length Number for each mRNA Background database Ensembl Gene ID Ensembl mRNA ID

8. OligoDB Output mRNA position T m of hybrid Probe stability Background matches Oligo probe

9. Biochemical Pathway Visualization Tools BioCarta EXPASY KEGG WIT MetaCyc

10. BioCarta Main focus: Pathways and tools to study them. Pathway tool is web-based. http://www.biocarta.com/genes/index.asp http://www.biocarta.com/genes/index.asp Static pages of pathways.  Metabolic and regulatory pathways. Need licence to add/edit pathways.

11. BioCarta Homepage

12. BioCarta Metabolic Pathway Example Glycolysis. Compounds shown in chemical and textual form. Enzymes above arrows with legend at bottom. Only enzymes are clickable.

13. BioCarta Regulatory Pathway Example ATM signalling pathway. All genes are clickable.

14. ExPASy “Expert Protein Analysis System”. Main focus: Analysis of protein sequences and structures. Pathway tool is web-based. http://www.expasy.org/cgi-bin/search-biochem-index Static pages of pathways.  Metabolic and Regulatory pathways.  Scanned in version of the Boehringer Mannheim “Biochemical Pathways” map.  Map partitioned into 115 pieces.  Keywords matched against entries in map. Cannot add/edit pathways.

15. ExPASy Homepage

16. ExPASy Metabolic Pathway Example Glycolysis. Color convention based on Michal’s “Biochemical Pathways”.  Color of compounds depends on compound type.  Color of arrows depend on species (plant, animal, yeast, prokaryotes). Only enzymes and arrows outside diagram are clickable.

17. ExPASy Regulatory Pathway Example Insulin receptors. Only enzymes and arrows outside diagram are clickable.

18. KEGG “Kyoto Encyclopedia of Genes and Genomes”. Main focus: Biochemical pathway visualization. Pathway tool is web-based. http://www.kegg.com/kegg/kegg2.html Static pages of pathways.  Metabolic and Regulatory pathways. Different organisms.

19. KEGG Homepage

20. KEGG Metabolic Pathway Example Glycolysis. Substrates and products are drawn in circles. Enzymes, represented by EC number, are drawn in rectangles. Adjacent pathways are drawn in semi-rectangles. Enzymes found in the gene catalog of a specific organism are marked green. All items are clickable.

21. KEGG Metabolic Pathway Example Select specific species Link to other pathway Link to compound Link to enzyme

22. KEGG Regulatory Pathway Example MAPK signaling pathway (Homo Sapiens).

23. WIT “What is There”. Main focus: Produce metabolic reconstructions for sequenced (or partially sequenced) genomes. Pathway tool is Web-based. http://wit.mcs.anl.gov/WIT2/ Static pages of pathways.  Only metabolic pathways.

24. WIT Homepage

25. WIT Metabolic Pathway Example Glycolysis. Enzymes. Substrates, and products are clickable.

26. MetaCyc Electronic encyclopedia of over 450 metabolic pathways from over 150 different organisms. Software:  Implemented in COMMON LISP; AI techniques incorporated to allow inference, such as pathway prediction from sequenced genomes.  Need a licence to obtain software.  Free for academic purposes only. Graph layout algorithm for drawing metabolic pathways dynamically at run-time. Need to manipulate database to add/edit pathways.

27. MetaCyc Software

28. MetaCyc Metabolic Pathway Example Glycolysis. Everything (Substrates, products, adjacent pathways, and arrows) is clickable.

29. Question Do the enzymes form the same pathway share a common regulation? Inspect Lysine Biosynthesis pathway. Use the following tools:  KEGG: http://www.genome.jp/kegg/http://www.genome.jp/kegg/  UCSC Genome Browser: genome.ucsc.edugenome.ucsc.edu  TransFac database or alternative search tools like TESS: http://www.cbil.upenn.edu/tess/http://www.cbil.upenn.edu/tess/