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flowchart TD
Omics([Omics technologies])
HTS[HTS]
MS[Mass spectrometry]
Illumina[Illumina]
ONT[ONT]
PacBio[PacBio]
Omics --> HTS
Omics --> MS
HTS --> Illumina
HTS --> ONT
HTS --> PacBio
classDef omics fill:#bb0000,color:#ffffff,stroke:#bb0000,font-weight:bold;
classDef hts fill:#009E73,color:#ffffff,stroke:#009E73,font-weight:bold;
classDef illumina fill:#0072B2,color:#ffffff,stroke:#0072B2,font-weight:bold;
classDef dim fill:#eaeaea,color:#999999,stroke:#cccccc;
class Omics omics;
class HTS hts;
class Illumina illumina;
class MS,ONT,PacBio dim;
The main omics data types
Copyright ThermoFisher
Learn more in this week’s first reading
Poinsignon et al. (2023) — Working with omics data:
An interdisciplinary challenge at the crossroads of biology and computer science
Figure 2 from the paper.
Libraries and library prep
In a sequencing context, a “library” is a collection of
nucleic acid fragments ready for sequencing.
These fragments number in the millions or billions and are
often simply generated at random from input such as genomic DNA:
An overview of the library prep procedure. This is typically done for you by a sequencing facility or company.
A closer look at the processed DNA fragments
As shown on the previous slide, after library prep,
each DNA fragment is flanked by several types of short sequences
that together make up the “adapters”:
Multiplexing!
Adapters can include so-called “indices” or “barcodes” that identify individual samples.
That way, many samples can be combined (multiplexed) into a single library.
Paired-end vs. single-end sequencing
DNA fragments can be sequenced from both ends as shown below,
and this is called “paired-end” (PE) sequencing:
Why paired-end sequencing?
This is a way for Illumina sequencing to produce more information from the same DNA fragment, given the limited read length of the technology (as you always know which two reads form a pair).
Paired-end vs. single-end sequencing
DNA fragments can be sequenced from both ends as shown below,
and this is called “paired-end” (PE) sequencing:
When sequencing is instead single-end (SE), no reverse read is produced:
Insert size variation
The DNA fragment’s size (“insert size”) can vary — by design, but also
because of limited precision in size selection. What happens when it is:
1: Shorter than the combined read length (but longer than a single read)?
- The fragment is too short to sequence, so it drops out of the library.
- The forward and reverse reads overlap in the middle.
- The reads come out shorter than the read length that was ordered.
- The final bases of each read consist of adapter sequence.
Insert size variation
The DNA fragment’s size (“insert size”) can vary — by design, but also
because of limited precision in size selection. What happens when it is:
2: Shorter than a single read? (The reads still overlap — but what else?)
- The sequencer stops early, so these reads are shorter than the rest.
- The reverse read is dropped, leaving only a forward read.
- The final bases of each read consist of adapter sequence.
- The fragment gets sequenced twice, producing a PCR duplicate.
Garrigós et al. 2025
Throughout the course, we’ll use an example/practice data set from Garrigós et al. (2025):
This paper uses paired-end Illumina RNA-Seq data to study gene expression in Culex pipiens mosquitoes infected with two different malaria-causing Plasmodium protozoans.