Technologies for whole genome sequencing and de novo assembly

# Technologies for whole genome sequencing and de novo assembly > Piotr Mieczkowski, UNC school of medicine **Short Reads**: Illumina, Ion torrent **Long Reads**: Pacific Bioscience, Oxford Nanopore **Optical Mapping**: Bionano Genomics Quantity application: RNAseq adn ChIPseq Qualitative: Whole genome assembly, ... de novo genome assembly ## Chromosome-level genome assembly ### 1. DNA extraction: most important thing is quality of material/DNA DNA extraction for optimal sequencing results **circulomics**: high yield, high molecular weigth DNA extraction (simple bind, wash and elute process). Very good if you want to extract large fragments ### 2. Illumina sequencing - short read is still more popular (cheaper) than long read sequencing - Illumina will soon be introducing long read technology (expensive but will be high quality and long reads around 10kb), these long read can be used for scaffolding - NovaSeq v1.5 reagents and key technology enables: *missed this part* Why is Illumina so popular? The chemistry 4 nucleotides, each contain a different fluorescent dye, add nucleotides, add them with dye, take picture, strip dye, add more nucleotides, repeat (some work based on 2 channel chemistry with only red and green dyes, each nucleotide has a different color, lack of color, or both) #### Hi-C (still short read) - helps you see chromosome folding - different types of Hi-C can serve as advanced chromatin confirmation technologies ### 3. Long Read Technologies (PacBio and Oxford Nanopore) - PacBio: Sequel II System - Single Molecule Real Time DNA seq (SMRT): technology allows DNA to loop so polymerase can go around the loop many types producing long, high-fidelity reads - read types: CLR and HiFI long reads - Oxford Nanopore - MinION - How Nanopore sequencing works: DNA goes through pour with an ion current. - read types: long and ultra long reads - you can get up to 250kb but important to have high quality DNA (high molecular weight DNA!!) What type of sequencing is best and what factors to consider: 1. Genome size Linearize DNA in NanoChannel array (??) Bionano Access software assembles de novo genome maps