Lecture 29: Analysing Microbial Genomes by NGS

1, Complex and advanced tech • Relies on: a. modern, high power computing b. Extensive engineering c. Nanoscale engineering d. Modern, advanced materials

• “Sequencing b synthesis” • Process: a. DNA amplified on a slide b. Fluorescent terminator base anneals c. Laser removes fluorescent terminator d. Fluorescent label read e. Process repeats

• Detection of fluorescence from labelled nucleotides during DNA synthesis • Occurs in arrays of small wells

• Long reads • Generating gold-standard reference genomes • Reconstruct intact genes and gene clusters • E.g., ANTIBIOTIC DISCOVERY

• Fast, high throughput, low cost/bp • Allows metagenomic analysis of complex environments • Can compare organisms cheaply

• If you want to understand how a gene is regulated. 2, Take something that glows or has specific output (LacZ blue/white, GFP) • Compare control with sample of KO TF

• Microarrays, qRT-PCR rely on hybridisation

\+ All transcription is studied in an unbiased manner \+ more precise - higher resolution \+ No interference from non-specific hybridization \+ Allows discovery of novel genetic features, delineation of operons and untranslated regions \+ greater dynamic range, more discrimitory and sensitive

• Identification of all the binding sites for a DNA binding proteins in a genome