11/06/2014 RNA  Seq   analysis   Cleaning   Plateforme  ABiMS RNA  Seq  analysis  :  cleanning Data  Cleaning Why  do  we  care  about  cleaning  ? Why  do  we  care  about  cleaning  ?   RAW  SEQUENCES Why  do  we  care  about  cleaning  ?   RAW  SEQUENCES   AMAZING TRANSCRIPTOME !!! Why  do  we  care  about  cleaning  ?   RAW  SEQUENCES   AMAZING TRANSCRIPTOME !!! NO !! Because…   •  Unknown  nucleoNdes   •  Bad  quality  nucleoNdes   •  Adaptors  and  primers  sub-­‐ sequences   •  Poly  A/T  tails   •  Low  complexity  sequences   •  rRNA  sequences   •  Contaminant  sequences   •  Short  length  sequences     But  also:   •  Removing  singletons   •  In-­‐silico  normalizaNon   •  Sequencing  errors  correcNon   •  … But  first…  What  data  do  we  have  ? NGS  sequences   •  Illumina,  454  (Roche),  Ion  Torrent,  Solid,  …   •  Single,  Paired-­‐end,  Mate  pairs   •  Sequences  length:  25,  35,  50,  75,  100,  150,   250,  500,  700,  800,  …  base  pairs   •  File  format:  Fastq  Phred+33,  Fastq  Phred+64,   2  files  (.fasta  +  .qual),  Colorspace