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Platforms available and what is best for my experiment?
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Short read sequencing is high throughput and can generate in many millions of reads. These reads are usually between 150-300bp long and have a high base accuracy
Long read sequencing is lower throughput in terms of the number of reads we see compared to short read sequencing. However the reads are many kbs in length
If in doubt about design, ask your facility you are using
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Understanding experimental design
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Most experiments need positive and negative controls
Technical replicates ensure our measures are reproducible
Biological replicates ensure our results generalise
Coverage is the percentage of the reference genome sequenced
Depth is the average number of times a base is sequenced
There is a trade-off between sequencing depth and the need for controls and replicates
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Statistical Analysis
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Type I errors are those caused by False positives.
Type II errors are those caused by False negatives.
We can reduce Type I errors by using a more stringent alpha threshold.
We can reduce Type II errors by improving the power in an experiment by increasing the number of replicates used.
High throughput experiments require us to use multiple testing correction. There are two popular methods to do this, the most stringent is the Bonferroni correction, the least stringent is the Benjamini Hochberg FDR.
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