Rapid genome sequencing with short universal tiling probes.
The increasing availability of high-quality reference genomic sequences has created a demand for ways to survey the sequence differences present in individual genomes. Here we describe a DNA sequencing method based on hybridization of a universal panel of tiling probes. Millions of shotgun fragments are amplified in situ and subjected to sequential hybridization with short fluorescent probes. Long fragments of 200 bp facilitate unique placement even in large genomes. The sequencing chemistry is simple, enzyme-free and consumes only dilute solutions of the probes, resulting in reduced sequencing cost and substantially increased speed. A prototype instrument based on commonly available equipment was used to resequence the Bacteriophage lambda and Escherichia coli genomes to better than 99.93% accuracy with a raw throughput of 320 Mbp/day, albeit with a significant number of small gaps attributed to losses in sample preparation.
Pihlak
Arno
A
Laboratory for Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Scheeles väg 1, SE-171 77 Stockholm, Sweden.
Baurén
Göran
G
Hersoug
Ellef
E
Lönnerberg
Peter
P
Metsis
Ats
A
Linnarsson
Sten
S
D002874:Chromosome Mapping / Q000639:trends*; D015342:DNA Probes / Q000235:genetics*; D005544:Forecasting; D017404:In Situ Hybridization, Fluorescence / Q000639:trends*; D017422:Sequence Analysis, DNA / Q000379:methods*; D013673:Technology Assessment, Biomedical
D015342:DNA Probes