UNIT 5

Question 1

Metagenomics

Answer 1

• Metagenomics
  
  • Study of genetic material from environmental samples
• Workflow
  
  • Sample collection and DNA extraction
  • Sequencing
  • Preprocessing and quality control
  • Taxonomic profiling and classification
  • Functional annotation
  • Assembly (optional)
  • Data visualization and interpretation
  • Statistical analysis
• Tools
  
  • QIIME/QIIME2, Kraken2, MEGAN, MetaPhlAn
  • FastQC, Trimmomatic, Cutadapt
  • Prokka, MetaGeneMark
• Applications
  
  • Human health
  • Environmental studies
  • Biotechnology
  • Agriculture
• Conclusion
  
  • Powerful tool for studying microbial communities
  • Provides insights into diversity, function, and interactions

Question 2

Metagenomics Raw Data Preprocessing

Answer 2

• Metagenomics Preprocessing
  
  • Cleaning and preparing raw sequencing data
• Steps
  
  • Quality assessment (FastQC)
  • Trimming and filtering (Trimmomatic, Cutadapt)
  • Adapter and contaminant removal (Cutadapt, BBMap, Decontam, KneadData)
  • Dereplication (USEARCH, VSEARCH)
  • Host DNA removal (Bowtie2, BBMap)
  • Merging paired-end reads (PEAR, Fastq-join)
  • Normalization (DESeq2, rarefaction)
  • Format conversion (SEQTK, FASTX-Toolkit)
• Importance
  
  • Improves data quality and accuracy
  • Removes errors and biases
  • Ensures reliable downstream analysis

Question 3

Metagenomic Features

Answer 3

• Metagenomic Features
  
  • Characteristics derived from metagenomic data
• Taxonomic Features
  
  • OTUs, ASVs
  • Taxonomic classification
  • Phylogenetic analysis
• Functional Features
  
  • Gene function prediction
  • Functional annotation
  • Metabolic pathway reconstruction
  • ARGs, virulence factors
• Diversity Metrics
  
  • Alpha diversity (Shannon, Chao1)
  • Beta diversity (Bray-Curtis, UniFrac)
  • Rarefaction curves
• Other Features
  
  • Co-occurrence networks
  • MAGs
  • Phage and viral identification
• Conclusion
  
  • Provide insights into microbial communities
  • Essential for understanding diversity, function, and interactions

Question 4

Metagenomic Taxonomy Assignment

Answer 4

• Taxonomy Assignment
  
  • Classifying metagenomic sequences into taxonomic groups
• Methods
  
  • Marker-gene based (16S/18S rRNA, ITS)
  • Shotgun metagenomics
• Tools
  
  • Kraken, MetaPhlAn, Centrifuge
  • QIIME, FastTree
  • BLAST, UCLUST, VSEARCH
• Challenges
  
  • Incomplete reference databases
  • Low resolution
  • Chimeric reads
  • Short reads
• Conclusion
  
  • Essential for understanding microbial communities
  • Tools and methods are continuously improving

Question 5

Phylogenetic Trees

Answer 5

• Phylogenetic Trees
  
  • Visualizing evolutionary relationships
• Key Components
  
  • Taxa
  • Branches
  • Nodes
  • Root
  • Clades
• Building Trees
  
  • Choosing data (DNA, protein, morphological)
  • Aligning sequences
  • Calculating distances
  • Building the tree (distance-based, character-based)
  • Evaluating the tree
• Importance
  
  • Understanding evolutionary history
  • Tracing disease evolution
  • Identifying new species
  • Making predictions about future evolution
• Conclusion
  
  • Essential tool for studying life on Earth