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Moderní metody analýzy genomu
- analýza
Mgr. Nikola Tom
Brno, 2.4.2014

Workflow
•


Raw sequence = fastq
•Biological sequence
•Corresponding quality scores
•ASCII character
•(fasta+ qual, csfasta + csqual, sff)
•
•@
•SEQ_ID GATTTGGGGTTCAAAGCAGTATCGATCAAATAGTAAATCCATTTGTTCAACTCACAGTTT
•+
• !''*((((***+))%%%++)(%%%%).1***-+*''))**55CCF>>>>>>CCCCCCC65

FastQC
•


Cutadapt
•Adaptor trimming (miRNA)
•Quality filtering
•Length filtering
•

•



Read mapping => SAM, BAM
•Usually mapping reads on reference
•miRNA - special case
–Grouping and annotate against mirBase
•DNA
–BWA, Bowtie, Bfast, SHRiMPclc
•RNA
–TopHat (de novo splice aligner)
•Commercial
–CLC Genomics Workbench

SAM
•


Alignment
•


Mapping, Coverage reports
•Important checkout for lab protocol
•Specificity of PCR
•Normalization
•Settings of variant calling threshold, CNV
•




SNV and small InDel Calling
•Coverage
•Frequency
•Base quality
•
•!!!
•Genomic context (homopolymers)
•Nucleotide type
•Position in read (errors at the read end)
•Alignment errors

CNV variations
•


Structural variations
•Mate-pair library
•Long InDel
•Translocation
•




Annotating and filtering
•Gene
•Transcript
•dbSNP
•Regulation
•Comparative genomics
•Repeats
•Functional
•Gene ontology
•miRNA targets
•Etc.