Index-free de novo assembly and deconvolution of mixed mitochondrial genomes.
Second-generation sequencing technology has allowed a very large increase in sequencing throughput. In order to make use of this high throughput, we have developed a pipeline for sequencing and de novo assembly of multiple mitochondrial genomes without the costs of indexing. Simulation studies on a mixture of diverse animal mitochondrial genomes showed that mitochondrial genomes could be reassembled from a high coverage of short (35 nt) reads, such as those generated by a second-generation Illumina Genome Analyzer. We then assessed this experimentally with long-range polymerase chain reaction products from mitochondria of a human, a rat, a bird, a frog, an insect, and a mollusc. Comparison with reference genomes was used for deconvolution of the assembled contigs rather than for mapping of sequence reads. As proof of concept, we report the complete mollusc mitochondrial genome of an olive shell (Amalda northlandica). It has a very unusual putative control region, which contains a structure that would probably only be detectable by next-generation sequencing. The general approach has considerable potential, especially when combined with indexed sequencing of different groups of genomes.
McComish
Bennet J
BJ
Allan Wilson Centre for Molecular Ecology and Evolution, Massey University, Palmerston North, New Zealand. b.mccomish@massey.ac.nz
Hills
Simon F K
SF
Biggs
Patrick J
PJ
Penny
David
D
D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D003198:Computer Simulation; D020451:Contig Mapping; D017931:DNA Primers / Q000235:genetics; D004272:DNA, Mitochondrial / Q000737:chemistry / Q000235:genetics*; D019143:Evolution, Molecular; D049851:Gastropoda / Q000235:genetics; D015894:Genome, Human; D054629:Genome, Mitochondrial; D006801:Humans; D019901:Locus Control Region; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D051381:Rats; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA / Q000379:methods* / Q000706:statistics & numerical data
D017931:DNA Primers; D004272:DNA, Mitochondrial