Abstract
Helicos™ Single Molecule Sequencing (SMS) provides a unique view of genome biology through direct sequencing of cellular nucleic acids in an unbiased manner, providing both accurate quantitation and sequence information. Sample preparation does not require ligation or PCR amplification, avoiding the GC-content and size biases observed in other technologies. DNA is simply sheared, tailed with poly(A), and hybridized to a flow cell surface containing oligo(dT) for sequencing-by-synthesis of billions of molecules in parallel. This process also requires far less material than other technologies. Gene expression measurements can be done using first-strand cDNA-based methods (RNA-Seq) or using a novel approach that allows direct hybridization and sequencing of cellular RNA for the most direct quantitation possible. In this unit, principles and methods for using the Helicos® Genetic Analysis System are discussed.