A friendly reminder on what measuring DNA by NanoDrop (a.k.a. random number generator) can actually tell youπ
Pure DNA has an A260/280 of ~1.8, while pure RNA has an A260/280 of ~2.0.
Pure DNA usually has an A260/230 of 2.0-2.2, although it can vary.
For DNA, spectrophotometers are good assessing purity, while fluorometers are good for assessing concentration.
Pure DNA usually has an A260/230 of 2.0-2.2, although it can vary.
For DNA, spectrophotometers are good assessing purity, while fluorometers are good for assessing concentration.
NanoDrop will often over-estimate concentration, especially when DNA is not perfectly pure.
It also loses reliability when DNA concentration is below 20 ng/uL.
NEB has a great technical note on the effect of many common contaminants.
tinyurl.com
It also loses reliability when DNA concentration is below 20 ng/uL.
NEB has a great technical note on the effect of many common contaminants.
tinyurl.com
Both concentration and quality are important for sequencing (especially nanopore sequencing).
Ideally, both a spectrophotometer and fluorometer should be used before to ensure DNA is pure and properly diluted.
Ideally, both a spectrophotometer and fluorometer should be used before to ensure DNA is pure and properly diluted.
Donβt believe the concentration reported by NanoDrop unless you are confident your DNA is absolutely pure.
Even then, itβs always best to verify concentration by fluorometer if you will be sequencing.
Even then, itβs always best to verify concentration by fluorometer if you will be sequencing.
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