  Background: The short reads output by first- and second-generation DNA
sequencing instruments cannot completely reconstruct microbial chromosomes.
Therefore, most genomes have been left unfinished due to the significant
resources required to manually close gaps in draft assemblies.
Third-generation, single-molecule sequencing addresses this problem by greatly
increasing sequencing read length, which simplifies the assembly problem.
  Results: To measure the benefit of single-molecule sequencing on microbial
genome assembly, we sequenced and assembled the genomes of six bacteria and
analyzed the repeat complexity of 2,267 complete bacteria and archaea. Our
results indicate that the majority of known bacterial and archaeal genomes can
be assembled without gaps, at finished-grade quality, using a single PacBio RS
sequencing library. These single-library assemblies are also more accurate than
typical short-read assemblies and hybrid assemblies of short and long reads.
  Conclusions: Automated assembly of long, single-molecule sequencing data
reduces the cost of microbial finishing to $1,000 for most genomes, and future
advances in this technology are expected to drive the cost lower. This is
expected to increase the number of completed genomes, improve the quality of
microbial genome databases, and enable high-fidelity, population-scale studies
of pan-genomes and chromosomal organization.
