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An Optimized Chloroplast DNA Extraction Protocol for Grasses (Poaceae) Proves Suitable for Whole Plastid Genome Sequencing and SNP Detection

Authors: Diekmann, Kerstin; Hodkinson, Trevor R; Fricke, Evelyn; Barth, Susanne;

An Optimized Chloroplast DNA Extraction Protocol for Grasses (Poaceae) Proves Suitable for Whole Plastid Genome Sequencing and SNP Detection

Abstract

Obtaining chloroplast genome sequences is important to increase the knowledge about the fundamental biology of plastids, to understand evolutionary and ecological processes in the evolution of plants, to develop biotechnological applications (e.g. plastid engineering) and to improve the efficiency of breeding schemes. Extraction of pure chloroplast DNA is required for efficient sequencing of chloroplast genomes. Unfortunately, most protocols for extracting chloroplast DNA were developed for eudicots and do not produce sufficiently pure yields for a shotgun sequencing approach of whole plastid genomes from the monocot grasses.We have developed a simple and inexpensive method to obtain chloroplast DNA from grass species by modifying and extending protocols optimized for the use in eudicots. Many protocols for extracting chloroplast DNA require an ultracentrifugation step to efficiently separate chloroplast DNA from nuclear DNA. The developed method uses two more centrifugation steps than previously reported protocols and does not require an ultracentrifuge.The described method delivered chloroplast DNA of very high quality from two grass species belonging to highly different taxonomic subfamilies within the grass family (Lolium perenne, Pooideae; Miscanthus x giganteus, Panicoideae). The DNA from Lolium perenne was used for whole chloroplast genome sequencing and detection of SNPs. The sequence is publicly available on EMBL/GenBank.

Keywords

570, Chloroplasts, DNA, Plant, Science, Genome, Plastid, Molecular Sequence Data, Genes, Plant, Poaceae, Polymorphism, Single Nucleotide, Smart & Sustainable Planet, Plant Genomics, Grasses, Plastids, Phylogeny, 580, Base Sequence, Q, R, DNA, Chloroplast, Chromosome Mapping, Sequence Analysis, DNA, Genetic Techniques, Medicine, Genes & Society, Genome, Plant, Research Article

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citations
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
46
Top 10%
Top 10%
Top 10%
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