The course aim to provide the basic elements for the analysis of “omic” data obtained by high-throughput sequencing and Proteomics in microbiology.
Analysis of NGS data and proteomics in bacteriology.
Bacterial Genomics: NGS sequencing methods and file formats. De-multiplexing, Sequence Quality Control (FastQC) and Read Trimmming. Strategies and software for read mapping, for de-novo assembly, samtools, Re-sequencing and variant calling from .bam files. Gene prediction and Genome annotation of bacterial genomes. On-line Bioinformatic tools for the analysis of bacterial genomes. Species identification, Virulome and acquired Resistome -Single Nucleotide Polymorphisms (SNPs) and acquired genes-. Detection and validation of specific SNPs by real time qPCR and High Resolution Meelting Analysis. Determination of in silico-MLST and SNP phylogenetic analysis. In silico MLST determination, genomic epidemiology and phylogeny by analysis of whole genome SNPs e core SNPs.
Bacterial RNAomes and Transcriptomics: Bacterial RNAomes and transcriptome analysis from RNA-seq raw data, read Trimming, de-novo transcript assembly and reference-genome transcript assembly. Normalization of data obtained from different experiments, analysis of differentially expressed genes and small non coding RNAs, determination of operon structures using bacterial transcriptome dedicated tools, Enrichment analysis and tools. Statistical data analysis by EdgeR or other software. Data validation by real time qPCR. Viewing results in IGV genome browser.
Gene Ontology: KEGG database, Go-consortium tools, analysis "functional protein association networks" by STRING Consortium tool.
Microbiomes: Analysis of Targeted Amplicon-Based Next Generation Sequencing of 16S rDNA and ITS for the study of bacterial microbiomes by the generation of OTU, descriptive (alpha-diversity and beta-diversity) and statistical analysis of microbiomes (QIIME). and Overview of Metagenomics.
Bacterial Proteomics: Spatial, temporal and environmental Proteomes. Methods for the proteome study (Two-dimensional electrophoresis (2-DE): IEF Separation by pI (isoelectric point), SDS-PAGE (separation by molecular mass), Image analysis (scanner), Mass spectrometry, Charge / mass ratios, Protein databases: EMBL, GenBank, SWISS PROT.
Metagenomics and Metatranscriptomics: Basics.