Sample Preparation, Sequencing Technology & NECs¶
Where the data comes from before it ever reaches QIIME2: sample collection, DNA extraction, library prep, the choice of sequencing platform, and the controls (positive, negative, mock) you should run alongside every batch.
What this lecture covers¶
- Sample collection and storage trade-offs (preservation buffers, freezing, chain-of-custody)
- DNA extraction kits, when the kit choice affects the answer
- 16S library prep: PCR primers, indexing, normalization
- Illumina chemistry vs alternatives, read length, error profile, and what that implies for ASVs
- NECs (negative extraction controls), positive controls, and mock communities, what each one tells you and what to do when they look bad
- A pre-flight checklist before you ever sequence
Hands-on follow-along¶
This block is lecture-only. Quality-control of sequencing controls is hands-on in Day 2, Quality Control.