Table 4 Overview of the special features of Sanger and Nanopore sequencing and their significance for practical work with both methods
Sanger | MinION Nanopore | |
|---|---|---|
Required DNA amount | 500 ng | 3–5 µg |
Required DNA quality | No special requirements | High Molecular Weight DNA |
Max. load of samples per run | 2 × 96 | 5 |
Amount of readable targets per sample (multiplexing) | 1 | Theoretically 20, but practically less, as the number of reads is crucial for the correct interpretation of the methylation data but is partly reduced by combining several sgRNAs. |
Reliability | Depending on the target and maybe methylation level. A good quality of sequencing (as indicated by high base Quality Values) is prerequisite for interpretation of data. | Supposedly high. Sufficient numbers of calls per site are prerequisite for interpretation of methylation data. |
Costs | Approximately 10,- €/sample If working in triplicates: 30,-€/sample | Approximately 150,- €/sample |
Preferably use for: | -Large patient cohorts -One target of interest -Relative alterations of methylation status between groups | -Smaller number of samples -Multiplexing -Absolute methylation values of single/individual patients -Simultaneous gain of sequence information like polymorphisms in the same strand -To check Sanger sequencing strategies before cohort analysis |