In this work, we describe Unique Molecular Identifiers (UMIs) that can be used to count absolute number of original molecules even after DNA amplification.
It is very difficult to detect individual DNA molecules in a complex mixture. Therefore, the signal is usually first amplified, making many copies of each molecule. Unfortunately, the copying complicates tracking the exact number of original molecules. The reason is that it is virtually impossible to tell afterwards exactly how many times each original molecule was copied as all copies originating from same type of molecules are indistinguishable from each other. In this work, we present a method in which the original molecules are artificially made different from each other, in such a way that the copies made from the different original molecules can be later distinguished. We show that the UMI method can be used to count absolute numbers of molecules. We also apply the method to simultaneously counting thousands of different types of messenger RNA molecules present in cells. The new method proved to be more accurate than the one that has been commonly used for this task. Efficient and reliable counting of messenger RNA molecules is important because their abundances reveal which genes are active in the cells of interest.