A flexible and efficient template format for circular consensus sequencing and SNP detection

Nucleic Acids Res. 2010 Aug;38(15):e159. doi: 10.1093/nar/gkq543. Epub 2010 Jun 22.


A novel template design for single-molecule sequencing is introduced, a structure we refer to as a SMRTbell template. This structure consists of a double-stranded portion, containing the insert of interest, and a single-stranded hairpin loop on either end, which provides a site for primer binding. Structurally, this format resembles a linear double-stranded molecule, and yet it is topologically circular. When placed into a single-molecule sequencing reaction, the SMRTbell template format enables a consensus sequence to be obtained from multiple passes on a single molecule. Furthermore, this consensus sequence is obtained from both the sense and antisense strands of the insert region. In this article, we present a universal method for constructing these templates, as well as an application of their use. We demonstrate the generation of high-quality consensus accuracy from single molecules, as well as the use of SMRTbell templates in the identification of rare sequence variants.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Base Sequence
  • Consensus Sequence
  • DNA / chemistry*
  • Oligonucleotides / chemistry*
  • Polymorphism, Single Nucleotide*
  • Sequence Analysis, DNA / methods*
  • Staphylococcus aureus / genetics
  • Templates, Genetic


  • Oligonucleotides
  • DNA