- Poster presentation
- Open Access
A cost-effective and universal strategy for complete prokaryotic genome sequencing proposed by computer simulation
Genome Biologyvolume 12, Article number: P6 (2011)
Pyrosequencing techniques allow scientists to perform prokaryotic genome sequencing and achieve draft sequences within a few days. However, the sequencing results always turn out to contain several hundred contigs. A multiplex PCR procedure is then needed to fill all of the gaps and to link the contigs into one full-length genome sequence [1–10]. The full-length prokaryotic genome sequence is the gold standard for comparative prokaryotic genome analysis. This study assessed pyrosequencing strategies by using a simulation with 100 prokaryotic genomes.
Our simulation shows the following: first, a single-end 454 Jr Titanium run combined with a paired-end 454 Jr Titanium run may assemble about 90% of 100 genomes into <10 scaffolds and 95% of 100 genomes into <150 contigs; second, the average contig N50 size is more than 331 kb (Table 1); third, the average single base accuracy is >99.99% (Table 1); fourth, the average false gene duplication rate is <0.7% (Table 1); fifth, the average false gene loss rate is <0.4% (Table 1); sixth, the total size of long repeats (both repeat length >300 bp and >700 bp) is significantly correlated to the number of contigs (Table 4); and, seventh, increasing the read length of a pyrosequencing run could improve the assembly quality significantly (Table 1, 2, 3).
A single-end 454 Jr run combined with a paired-end 454 Jr run is a good strategy for prokaryotic genome sequencing. This strategy provides a solution to producing a high-quality draft genome sequence of almost any prokaryotic organism, selected at random, within days. It could be the first step to achieving the full-length genome sequence. It also makes the subsequent multiplex PCR procedure (for gap filling) much easier, aided by the knowledge of the orders/orientations of most of the contigs. As a result, large-scale full-length prokaryotic genome-sequencing projects could be finished within weeks.
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