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      Chloroplast genomic resources for phylogeny and DNA barcoding: a case study on Fritillaria

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          Abstract

          The genus Fritillaria comprises approximately 130 perennial herbaceous species. In the Pharmacopoeia of the People’s Republic of China, the bulbs of 11 Fritillaria species are used in Chinese herbal medicines. However, the traditional methods of morphological classification cannot accurately identify closely related species of Fritillaria. Previous studies have attempted to identify these species with universal molecular markers, but insufficient phylogenetic signal was available. In this study, the complete chloroplast genomes of eight Fritillaria species were compared. The length of the eight Fritillaria chloroplast genomes ranges from 151,009 bp to 152,224 bp. A total of 136 SSR loci were identified, including 124 polymorphic SSR loci. For large repeat sequences, 108 repeat loci and four types of repeats were observed. Ten highly variable regions were identified as potential molecular markers. These SSRs, large repeat sequences and highly variable regions provide important information for the development of genetic markers and DNA fingerprints. Phylogenetic analyses showed that the topological structures of all data sets (except the IR regions) were in complete agreement and well resolved. Overall, this study provides comprehensive chloroplast genomic resources, which will be valuable for future studies of evolution and species identification in Fritillaria.

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          OrganellarGenomeDRAW—a suite of tools for generating physical maps of plastid and mitochondrial genomes and visualizing expression data sets

          Mitochondria and plastids (chloroplasts) are cell organelles of endosymbiotic origin that possess their own genetic information. Most organellar DNAs map as circular double-stranded genomes. Across the eukaryotic kingdom, organellar genomes display great size variation, ranging from ∼15 to 20 kb (the size of the mitochondrial genome in most animals) to >10 Mb (the size of the mitochondrial genome in some lineages of flowering plants). We have developed OrganellarGenomeDraw (OGDRAW), a suite of software tools that enable users to create high-quality visual representations of both circular and linear annotated genome sequences provided as GenBank files or accession numbers. Although all types of DNA sequences are accepted as input, the software has been specifically optimized to properly depict features of organellar genomes. A recent extension facilitates the plotting of quantitative gene expression data, such as transcript or protein abundance data, directly onto the genome map. OGDRAW has already become widely used and is available as a free web tool (http://ogdraw.mpimp-golm.mpg.de/). The core processing components can be downloaded as a Perl module, thus also allowing for convenient integration into custom processing pipelines.
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            Highly Variable Chloroplast Markers for Evaluating Plant Phylogeny at Low Taxonomic Levels and for DNA Barcoding

            Background At present, plant molecular systematics and DNA barcoding techniques rely heavily on the use of chloroplast gene sequences. Because of the relatively low evolutionary rates of chloroplast genes, there are very few choices suitable for molecular studies on angiosperms at low taxonomic levels, and for DNA barcoding of species. Methodology/Principal Findings We scanned the entire chloroplast genomes of 12 genera to search for highly variable regions. The sequence data of 9 genera were from GenBank and 3 genera were of our own. We identified nearly 5% of the most variable loci from all variable loci in the chloroplast genomes of each genus, and then selected 23 loci that were present in at least three genera. The 23 loci included 4 coding regions, 2 introns, and 17 intergenic spacers. Of the 23 loci, the most variable (in order from highest variability to lowest) were intergenic regions ycf1-a, trnK, rpl32-trnL, and trnH-psbA, followed by trnSUGA-trnGUCC , petA-psbJ, rps16-trnQ, ndhC-trnV, ycf1-b, ndhF, rpoB-trnC, psbE-petL, and rbcL-accD. Three loci, trnSUGA-trnGUCC , trnT-psbD, and trnW-psaJ, showed very high nucleotide diversity per site (π values) across three genera. Other loci may have strong potential for resolving phylogenetic and species identification problems at the species level. The loci accD-psaI, rbcL-accD, rpl32-trnL, rps16-trnQ, and ycf1 are absent from some genera. To amplify and sequence the highly variable loci identified in this study, we designed primers from their conserved flanking regions. We tested the applicability of the primers to amplify target sequences in eight species representing basal angiosperms, monocots, eudicots, rosids, and asterids, and confirmed that the primers amplified the desired sequences of these species. Significance/Conclusions Chloroplast genome sequences contain regions that are highly variable. Such regions are the first consideration when screening the suitable loci to resolve closely related species or genera in phylogenetic analyses, and for DNA barcoding.
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              A Phylogenetic Analysis of 34 Chloroplast Genomes Elucidates the Relationships between Wild and Domestic Species within the Genus Citrus

              Citrus genus includes some of the most important cultivated fruit trees worldwide. Despite being extensively studied because of its commercial relevance, the origin of cultivated citrus species and the history of its domestication still remain an open question. Here, we present a phylogenetic analysis of the chloroplast genomes of 34 citrus genotypes which constitutes the most comprehensive and detailed study to date on the evolution and variability of the genus Citrus. A statistical model was used to estimate divergence times between the major citrus groups. Additionally, a complete map of the variability across the genome of different citrus species was produced, including single nucleotide variants, heteroplasmic positions, indels (insertions and deletions), and large structural variants. The distribution of all these variants provided further independent support to the phylogeny obtained. An unexpected finding was the high level of heteroplasmy found in several of the analyzed genomes. The use of the complete chloroplast DNA not only paves the way for a better understanding of the phylogenetic relationships within the Citrus genus but also provides original insights into other elusive evolutionary processes, such as chloroplast inheritance, heteroplasmy, and gene selection.
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                Author and article information

                Contributors
                dyp_851212@126.com
                zhangxiuhai@baafs.net.cn
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                19 January 2018
                19 January 2018
                2018
                : 8
                : 1184
                Affiliations
                [1 ]ISNI 0000 0004 0646 9053, GRID grid.418260.9, Beijing Agro-Biotechnology Research Center, , Beijing Academy of Agriculture and Forestry Sciences, ; Beijing, 100097 P. R. China
                [2 ]Beijing Key Laboratory of Agricultural Genetic Resources and Biotechnology, Beijing Engineering Technology Research Center of Functional Floriculture, Beijing, 100097 P. R. China
                [3 ]Engineering Research Center of Mt. Changbai Ecological Resources Development, Changhcun Sci-Tech University, Changchun, 130600 Jilin Province P. R. China
                Article
                19591
                10.1038/s41598-018-19591-9
                5775360
                29352182
                f5d258c8-293a-4cd7-8443-4d691c49f156
                © The Author(s) 2018

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 14 September 2017
                : 4 January 2018
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