What is the “extra purification step” that is needed for many of the fluorescent dye oligonucleotide conjugates?

Dyes listed with the “extra purification step” requirement are made in a two-step process. First, the unlabeled oligonucleotide with the appropriate amino or thiol linker is synthesized and then purified to isolate only the full-length oligonucleotide. The oligonucleotide is then conjugated to the dye and purified again to remove any unlabeled material. Some fluorescent dyes, such as 6-FAM, TET and HEX, do not require this extra purification step. These dyes are available as phosphoramidite reagents, allowing us to label the oligonucleotide during the synthesis procedure and use a single purification step.