Safe-Sequencing System is a Unique Molecular Identifier (UMI) Approach to Detect Rare Variants

Safe-SeqS (or, more commonly, Safe-Seq) is an approach that uses unique molecular identifiers (UMIs) to detect rare variants (Kinde et al., 2011). In the time after the publication of the method in 2011, the use of UMIs has become ubiquitous, particularly in single-cell sequencing approaches, and the name of the method fell into disuse.

Safe-Seq assigns a UMI to each template molecule and amplifies each uniquely tagged template molecule to create UMI families. The abundance of each UMI can be used to distinguish between rare mutations and technical errors, as well as to correct for PCR amplification bias.


  • Distinguishes between rare mutations and technical errors
  • Detects 1 mutant template among 5000 to 1,000,000 wild-type templates (Kinde et al., 2013)
  • Corrects for PCR amplification bias


  • Can introduce spurious cross-hybridization
  • Complex protocol that requires a gel-purification step (Stahlberg et al., 2016)
  • Uses 2 or 4 PCR cycles for barcode addition and does not satisfy the basic principle of labeling each molecule with a single, unique barcode (Kukita et al., 2015)


Illumina Library prep and Array Kit Selector


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