Capturing chloroplast variation for molecular ecology studies: a simple next generation sequencing approach applied to a rainforest tree

McPherson, Hannah, van der Merwe, Marlien, Delaney, Sven K., Edwards, Mark A., Henry, Robert J., McIntosh, Emma, Rymer, Paul D., Milner, Melita L., Siow, Juelian and Rossetto, Maurizio (2013) Capturing chloroplast variation for molecular ecology studies: a simple next generation sequencing approach applied to a rainforest tree. BMC Ecology, 13 8.1-8.11. doi:10.1186/1472-6785-13-8


Author McPherson, Hannah
van der Merwe, Marlien
Delaney, Sven K.
Edwards, Mark A.
Henry, Robert J.
McIntosh, Emma
Rymer, Paul D.
Milner, Melita L.
Siow, Juelian
Rossetto, Maurizio
Title Capturing chloroplast variation for molecular ecology studies: a simple next generation sequencing approach applied to a rainforest tree
Journal name BMC Ecology   Check publisher's open access policy
ISSN 1472-6785
Publication date 2013-03
Year available 2013
Sub-type Article (original research)
DOI 10.1186/1472-6785-13-8
Open Access Status DOI
Volume 13
Start page 8.1
End page 8.11
Total pages 11
Place of publication London, United Kingdom
Publisher BioMed Central
Collection year 2014
Language eng
Formatted abstract
Background: With high quantity and quality data production and low cost, next generation sequencing has the potential to provide new opportunities for plant phylogeographic studies on single and multiple species. Here we present an approach for in silicio chloroplast DNA assembly and single nucleotide polymorphism detection from short-read shotgun sequencing. The approach is simple and effective and can be implemented using standard bioinformatic tools.

Results: The chloroplast genome of Toona ciliata (Meliaceae), 159,514 base pairs long, was assembled from shotgun sequencing on the Illumina platform using de novo assembly of contigs. To evaluate its practicality, value and quality, we compared the short read assembly with an assembly completed using 454 data obtained after chloroplast DNA isolation. Sanger sequence verifications indicated that the Illumina dataset outperformed the longer read 454 data. Pooling of several individuals during preparation of the shotgun library enabled detection of informative chloroplast SNP markers. Following validation, we used the identified SNPs for a preliminary phylogeographic study of T. ciliata in Australia and to confirm low diversity across the distribution.

Conclusions: Our approach provides a simple method for construction of whole chloroplast genomes from shotgun sequencing of whole genomic DNA using short-read data and no available closely related reference genome (e.g. from the same species or genus). The high coverage of Illumina sequence data also renders this method appropriate for multiplexing and SNP discovery and therefore a useful approach for landscape level studies of evolutionary ecology.

Keyword Illumina
Shotgun sequencing
Chloroplast genome
Snp
Phylogeography
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Article number 8.

Document type: Journal Article
Sub-type: Article (original research)
Collections: Queensland Alliance for Agriculture and Food Innovation
Official 2014 Collection
 
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