1. June 2013 Metagenomics of Marine Microbes Tutorial overview Daniel Vaulot Station Biologique de Roscoff

2. June 2013 Bioinformatics analysis Reads Contigs Known genome Assembly Coverage Haplotypes BLAST Phylogenetic Diversity Functional Diversity Annotation Phylogeny Synteny Functions Biogeography Bioedit Newbler Bowtie Tablet Geneious MEGAN blastall Camera Size, GC% R R Camera

3. June 2013 Three environments Windows desktop ABIMS cluser (Linux) Web based services

4. June 2013 Software to be used SoftwareAimOperating system BioeditVisualize sequenceWindows RCompute Length and GC%Windows/Linux CameraBLASTWeb MEGANAnalysis of BLAST resultsJava (Windows) NewblerAssembly de novoLinux BowtieAssembly to referenceLinux TabletVisualize assemblyJava (Windows) ODVVisualize oceanographic dataWindows GeneiousIntegrated software (demo only) Java (Windows)

5. June 2013 Where to find sofware Windows * Bioedit (http://www.mbio.ncsu.edu/bioedit/page2.html) * R with Bioconductor pacakge installed (http://www.r-project.org/) * Rstudio * PuTTY * WinSCP or FileZilla (ftp client) * MEGAN (http://ab.inf.uni-tuebingen.de/software/megan/) * Tablet (http://bioinf.scri.ac.uk/tablet/) * Ocean Data View (http://odv.awi.de/) Web * CAMERA : https://portal.camera.calit2.net/gridsphere/gridsphere?cid=home ABIMS cluster * BLAST * MEGAN * Newbler * Bowtie

6. June 2013 Reads files BIOSOPE_T142_reads_random.fasta BIOSOPE_T142_reads_Bathy.fasta

7. June 2013 Steps 1.Prepare the environments and transfer files 2.Inspection and basic characteristics of reads 3.BLAST 4.Analysis of BLAST results with MEGAN 5.Assembly de novo and to reference Genome 6.Searching other metagenomes

8. 2 – Inspect and Compute basic statistics - Bioedit

9. 2 – Inspect and Compute basic statistics - R

10. June 2013 3 – BLASTN - Camera

11. June 2013 Import data

12. June 2013 Camera workflows

13. June 2013 Launch workflow - BLASTn

14. June 2013 Worflow results Download as : - xls (Excel) - xml (Megan) Download as : - xls (Excel) - xml (Megan)

15. June 2013 Analyze results with Excel

16. June 2013 4 - Import xml file into MEGAN

17. June 2013 MEGAN with full data set BLASTN Nb of hits : 25 Output format XML Import into MEGAN

18. June 2013 5 - Assembly – de novo - CAMERA

19. June 2013 5 – Assemble de novo with Newbler

20. June 2013 Contigs Reads mito Reads random

21. June 2013 5 – Assemble to reference Reads mito

22. June 2013 Assemble to reference Bowtie (on the ABIMS cluster)

23. June 2013 Tablet viewer Bowtie – 216 reads assembled Geneious – 252 reads assembled

24. June 2013 Mutations (Variants)

25. June 2013 6 - Search for other metagenomics reads corresponding to Bathycoccus mitochondrial genome

26. June 2013 Metagenome reads

27. June 2013 Stations where Bathycoccus mitochondrial reads detected in CAMERA data set 1448 423 418 327 309 223 168

28. June 2013 Geneious An integrated solution

29. June 2013 Geneious

30. June 2013 Reads Geneious for metagenomics analyses

31. June 2013 Create contigs

32. June 2013 Assembly into contigs

33. June 2013 Genomes Explore with browser

34. June 2013 Assemble/Map to reference Bathycoccus chromosome 000

35. June 2013 Assemble/Map to reference Bathycoccus chromosome 000 Coverage

36. June 2013 Detect nucleotide variation

37. June 2013 Detect nucleotide variation