Displaying NGS data Osvaldo Graña ograna@cnio.es Madrid, Marzo 2012

1. Structural Biology and Biocomputing Programme Displaying NGS data Osvaldo Graña ograna@cnio.es Madrid, Marzo 2012

2. How to upload a BAM file to the browser? BAM files are quite big, we cannot upload these files to the browser. But we can create an index file for the BAM file, this way the genome browser is able to upload only the information inside the BAM file for the region that we are visualizing. Steps: 1)create the INDEX file for the BAM file (with Samtools) cd samtools-0.1.18/misc/ samtools index ../../163adh.bam ****A new file was created '163adh.bam.bai', the BAM index file 3)put these two files together in a computer accesible from outside scp 163adh.bam* ograna@ubio:public_html/cursos/Master_Bioinformatica_2013/UCSC/

3. How to upload a BAM file to the browser? 4)Now we can access the files through an internet browser

4. How to upload a BAM file to the browser? 5)now we can create the new track in the browser **** remember that the reads comes from a human tissue

5. How to upload a BAM file to the browser? * adding the new track for the BAM file

6. How to upload a BAM file to the browser? * DEFINING the new track for the BAM file

7. How to upload a BAM file to the browser? * DEFINING the new track for the BAM file

8. How to upload a BAM file to the browser? * the new track is successfully defined and ready to use if we go back to the browser

9. How to upload a BAM file to the browser? * the reads contained in the BAM file are now shown in the browser, paste the following position: chr10:73590778-73591952

10. How to create a BED file from a BAM file with Bedtools 1)convert BAM to BED (original reads in the BAM are 40bp length) ./BEDTools-Version-2.16.1/bin/bedtools bamtobed -i 163adh.bam > 163adh.bed 3)edit the bed file and add the BED header as the first line

11. Uploading the BED file to the genome browser 1.- Select the proper genome and assembly (the reads come from a human tissue)

12. Uploading the BED file to the genome browser * click 'add custom tracks'

13. Uploading the BED file to the genome browser * select the BED file

14. Uploading the BED file to the genome browser * The BED file is correctly uploaded, now we go back to the browser

15. Back in the browser

16. Back in the browser

17. How to change the BED file coordinates to the latest assembly mm10 *For this exercise we are going to create first a NEW bed file directly in the browser in the mouse mm9 assembly

18. How to change the BED file coordinates to the latest assembly mm10 *Create a new bed file DIRECTLY in the browser: select mouse and the mm9 assembly

19. How to change the BED file coordinates to the latest assembly mm10 *Now we paste the coordinates for 3 theoretical reads

20. How to change the bed file coordinates to the latest assembly mm10 *we send the coordinates to create the new track

21. How to change the bed file coordinates to the latest assembly mm10 *the new track has been created, now we go back to the browser

22. How to change the bed file coordinates to the latest assembly mm10 *The reads that we created are shown in the 3'UTR of the mouse Mdm1 gene

23. How to change the bed file coordinates to the latest assembly mm10 *The reads that we created are shown in the 3'UTR of the mouse Mdm1 gene THESE COORDINATES ARE NOT VALID for the mouse MM10 assembly, we are going to check it now

24. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *Rebuilding the track using mm10 with the coodinates of mm9

25. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *Rebuilding the track using mm10 with the coodinates of mm9

26. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *Rebuilding the track using mm10 with the coodinates of mm9

27. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *Rebuilding the track using mm10 with the coodinates of mm9

28. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *In the mm10 assembly they are not over the Mdm1 gene anymore because coordinates has changed. We have to translate the coordinates from mm9 to mm10.

29. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly * We have to use the liftOver tool

30. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly * We have to use the liftOver tool

31. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly * We have to use the liftOver tool

32. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly * We have to use the liftOver tool

33. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly * We have to use the liftOver tool

34. The same coordinates of the reads in mm9 do not work with the mouse mm10 assembly *Converted coordinates

35. New coordinates for mouse mm10 assembly *We add now the new mm10 custom track with the new coordinates

36. New coordinates for mouse mm10 assembly *We add now the new mm10 custom track with the new coordinates

37. New coordinates for mouse mm10 assembly We add now the new mm10 custom track with the new coordinates

38. New coordinates for mouse mm10 assembly *Now we have the same reads shown in the same region of gene Mdm1 but in the latest mouse assembly mm10

39. Don't loose your data in the UCSC genome browser, create a user account

40. Don't loose your data in the UCSC genome browser, create a user account

41. Don't loose your data in the UCSC genome browser, create a user account

42. Don't loose your data in the UCSC genome browser, create a user account