Application of gene sequences in plant phylogenetic inferences

Document Type : Review Paper


Department of Cell and Molecular Biology, Faculty of Chemistry, University of Kashan, P. O. Box: 873175-3153, Kashan, Iran.


Molecular phylogenetic is the branch of phylogeny that analyzes hereditary molecular diversity, mainly in DNA sequences, to increase data on an organism‘s evolutionary relationships. Due to the taxonomic levels of the study, various molecular markers are applied in molecular phylogeny. The selection of molecular instrument is of paramount matter to ensure that a proper level of variation is meliorated to respond the phylogenetic question. In this review, we have been trying to discuss about gene markers used in the plant phylogeny at various taxonomic levels. The current gene markers used in phylogeny include: the ribosomal nuclear genes, low copy nuclear genes and the extra-nuclear genome (mitochondrial and chloroplastic genomes). Conserved regions could be used at higher taxonomic levels in phylogenetics studies and regions with more changes could be applied between closely related taxa. One of the most common sequences for studying the phylogenetic relationships at the generic and infrageneric taxonomic levels in plants is the internal transcribed spacer (ITS) region of the 18S–5.8S–26S nuclear ribosomal cistron. Chloroplastic gene sequences have been used extensively at the family level and above but chloroplast non-coding sequences such as introns and intergenic spacers are used more frequently at lower taxonomic levels. Low-copy nuclear genes are most useful at the interspecific and intraspecific levels where cpDNA and/or nrDNA cannot provide adequate resolution. Evidence offers that for more strongly reconstruction of phylogeny, several discrete genes are needed. Now, uses of next generation sequencing (NGS) techniques are reported. Techniques for NGS are an alternative to prevalent methods that let access to hundreds of DNA regions.


Abreu N. L. d., Alves R. J. V., Cardoso S. R. S., Bertrand Y. J. K., Sousa F., Hall C. F., Pfeil B. E., and Antonelli A. (2018). The use of chloroplast genome sequences to solve phylogenetic incongruences in Polystachya Hook (Orchidaceae Juss). Peer-reviewed Journal, 6: e4916.
Almerekova S., Mukhitdinov N., and Abugalieva S. (2017). Phylogenetic study of the endemic species Oxytropis almaatensis (Fabaceae) based on nuclear ribosomal DNA ITS sequences. BMC Plant Biology, 17(1): 173.
Azani N., Bruneau A., Wojciechowski M. F., and Zarre S. (2017). Molecular phylogenetics of annual Astragalus (Fabaceae) and its systematic implications, Botanical Journal of the Linnean Society, 184 (3): 347-365.
Babineau M., Gagnon E., and Bruneau A. (2013). Phylogenetic utility of 19 low copy nuclear genes in closely related genera and species of caesalpinioid legumes. South African Journal of Botany, 89: 94-105.
Baldwin B .G., Sanderson M .J., Porter J .M., Wojciechowski M .F., Campbell C .S., and Donoghue M . J. (1995). The ITS region of nuclear ribosomal DNA: A valuable source of evidence on angiosperm phylogeny. Annals of the Missouri Botanical Garden, 82(2): 247-277.
Baldwin B. G. (1992). Phylogenetic utility of the internal transcribed spacers of nuclear ribosomal DNA in plants: An example from the Compositae. Molecular Phylogenetics and Evolution, 1(1): 3-16.
Bellarosa R., Simeonea M. C., Papinib A., and Schironea B. (2005). Utility of ITS sequence data for phylogenetic reconstruction of Italian Quercus spp. Molecular Phylogenetics and Evolution, 34: 355-370.
Bult C. J., and Zimmer E. A. (1993). Nuclear ribosomal RNA sequences for inferring tribal relationships within Onagraceae. Systematic Botany, 18: 48-63.
Bult C. J., Sweere J. A., and Zimmer E. A. (1995). Cryptic sequence simplicity, nucleotide composition bias, and molecular coevolution in the large subunit of ribosomal DNA in plants: implications for phylogenetic analyses. Annals of the Missouri Botanical Garden, 82: 235-246.
Carbonell-Caballero J., Alonso R., Ibanez V., Terol J., Talon M., and Dopazo J. (2015). A phylogenetic analysis of 34 chloroplast genomes elucidates the relationships between wild and domestic species within the genus Citrus. Molecular Biology and Evolution, 32(8): 2015-2035.
Cascales J., Bracco M., Garberoglio M. J., Poggio L., and Gottlieb A. M. (2017). Integral phylogenomic approach over Ilex L. species from Southern South America. Life, 47(7): 2-18.
CBOL Plant Wording Group. (2009). A DNA barcode for land plants. Proceedings of the National Academy of Sciences, 106: 12794–12797.
Cerros-Tlatilpa R., Columbus J. T., and Barker N. P. (2011). Phylogenetic relationships of Aristida and relatives (Poaceae, Aristidoideae) based on noncoding chloroplast (trnL-F, rpl16) and nuclear (ITS) DNA sequences. American Journal of Botany, 98(11): 86-1868.
Chery J. G., Sass C., and Specht C. D. (2017). Development of single-copy nuclear intron markers for species-level phylogenetics: case study with Paullinieae (Sapindaceae). Applications in Plant Sciences, 5(9): 1700051.
Choi K. S., Kwak M., Lee B., and Park S. (2018). Complete chloroplast genome of Tetragonia tetragonioides: Molecular phylogenetic relationships and evolution in Caryophyllales. PLOS One, 13(6): e0199626.
Cronn R. C., Small R. L., and Wendel J. F. (1999). Duplicated genes evolve independently after polyploid formation in cotton. Proceedings of the National Academy of Sciences of the United States of America, 96(25): 14406-14411.
Cronn R. C., Small R. L., Haselkorn T., and Wendel J. F. (2002). Rapid diversification of the cotton genus (Gossypium: Malvaceae) revealed by analysis of sixteen nuclear and chloroplast genes. American Journal of Botany, 89: 707-725.
Cronn R. C., Small R. L., Haselkorn T., and Wendel J. F. (2003). Cryptic repeated genomic recombination during speciation in Gossypium gossypioides. Evolution, 57(11): 2475-2489.
Dechbumroong P., Aumnouypol S., Denduangboripant J., and Sukrong S. (2018). DNA barcoding of Aristolochia plants and development of species-specific multiplex PCR to aid HPTLC in ascertainment of Aristolochia herbal materials. PLOS One, 13(8): e0202625.
Doh E. J., Paek S. H., Lee G., Lee M. Y., and Oh S. E. (2016). Application of partial jnternal transcribed spacer sequences for the discrimination of Artemisia capillaris from other Artemisia species. Evidence-based complementary and alternative medicine, 2016: 7043436.
Dong W. L., Wang R. N., Zhang N. Y., Fan W. B., Fang M. F., and Li Z. H. (2018). Molecular evolution of chloroplast genomes of Orchid species: insights into phylogenetic relationship and adaptive evolution. International Journal of Molecular Sciences, 19(3): 716.
Donoghue M. J., and Mathews S. (1998). Duplicate Genes and the root of angiosperms, with an example using phytochrome sequences. Molecular Phylogenetics and Evolution, 9(3): 489-500.
Fan C., and Xiang Q- Y. (2003). Phylogenetic analyses of Cornales based on 26S rRNA and combined 26S rDNA-MATK-RBCL sequence data. American Journal of Botany, 90 (9): 1357-1372.
Fan X., Liu J., Sha L. N., Sun G. L., Hu Z. Q., Zeng J., Kang H. Y., Zhang H. Q., Wang Y., Wang X. L., Zhang L., Ding C. B., Yang R. W., Zheng Y. L., and Zhou Y. H. (2014) Evolutionary pattern of rDNA following polyploidy in Leymus (Triticeae: Poaceae). Molecular Phylogenetics and Evolution, 77: 296-306.
Fan C. (2001). Phylogenetic relationships within Cornus (Cornaceae) based on 26S rDNA sequences. American journal of botany, 88 (6): 8-1131.
Fitch W. M. (1970). Distinguishing homologous from analogous proteins. Systematic Zoology, 19(2): 99-113.
Fitch W. M. (2000). Homology a personal view on some of the problems. Trends in Genetics, 16: 227-231.
Galloway G. L., Malmberg R. L., and Price R. A. (1998). Phylogenetic utility of the nuclear gene arginine decarboxylase: an example from Brassicaceae. Molecular Biology and Evolution, 15(10): 1312-1320.
Gao X., Zhu Y. P., Wu B. C., Zhao Y. M., Chen J. Q., and Hang Y. Y. (2008). Phylogeny of Dioscorea sect. Stenophora based on chloroplast matK, rbcL and trnL-F sequences. Journal of Systematics and Evolution, 46(3): 315-321.
Ghada B., Ahmed B. A., Messaoud M., and Amel S. H. (2013). Genetic diversity and molecular evolution of the internal transcribed spacer (ITSs) of nuclear ribosomal DNA in the Tunisian fig cultivars (Ficus carica L.; Moracea). Biochemical Systematics and Ecology, 48: 20-33.
Gillespie L. J., Soreng, Robert John, Bull R. D., Jacobs S. W. L., and Refulio-Rodriguez N. F. (2008). Phylogenetic relationships in subtribe Poinae (Poaceae, Poeae) based on nuclear ITS and plastid trnT-trnL-trnF sequences. Botany, 86 (8): 938-967.
Giudicelli G. C., Mäder G., Silva-Arias G. A., Zamberlan P. M., Bonatto S. L., and Freitas L. B. (2017). Secondary structure of nrDNA Internal Transcribed Spacers as a useful tool to align highly divergent species in phylogenetic studies. Genetics and Molecular Biology, 40 (1): 191-199.
Hamby K. R., and Zimmer E. A. (1988). Ribosomal RNA sequences for inferring phylogeny within the grass family (Poaceae). Plant Systematics and Evolution, 160: 29-37.
Hamby K. R., and Zimmer E. A. (1992). Ribosomal RNA as a phylogenetic tool in plantsystematics. In Soltis P., Soltis D., and Doyle J. (Eds.), Molecular systematics of plants, New York: Chapman & Hall, 50- 91.
Hawkins J. S., Ramachandran D., Henderson A., Freeman J., Carlise M., Harris A., and Willison-Headley Z. (2015). Phylogenetic reconstruction using four low-copy nuclear loci strongly supports a polyphyletic origin of the genus Sorghum. Annals of Botany, 116(2): 291-299.
Hershkovitz M. A., and Lewis L. A. (1996). Deep-level diagnostic value of the rDNA-ITS region. Molecular Biology and Evolution, 13(9): 76-95.
Hershkovitz M. A., Zimmer E. A., and Hohn W. J. (1999). Ribosomal DNA sequences and angiosperm systematics. In Hollingsworth P. M., Bateman R. M., and Gornall R. J. (Eds.), Molecular systematics and plant evolution, London: Taylor & Francis, 268-326.
Hiesel R., Haeseler A. V., and Brennicke A. (1994). Plant mitochondrial nucleic acid sequences as a tool for phylogenetic analysis. Proceedings of the National Academy of Sciences of the United States of America, 91(2): 634-638.
Hilu K. W., Black C., Diouf D., and Burleigh J. G. (2008). Phylogenetic signal in matK vs. trnK: A case study in early diverging Eudicots (Angiosperms). Molecular Phylogenetics and Evolution, 48(3): 1120-1130.
Hollingsworth P. M., Graham S. W., and Little D. P. (2011). Choosing and using a plant DNA barcode. PLoS One, 6: e19254.
Ishikawa N., Yokoyama J. and Tsukaya H. (2009). Molecular evidence of reticulate evolution in the subgenus Plantago (Plantaginaceae). American Journal of Botany, 96(9): 1627-1635.
Jansen R. K., and Palmer J. D. A. (1987). Chloroplast DNA inversion marks an ancient evolutionary split in the sunflower family (Asteraceae). Proceedings of the National Academy of Sciences of the United States of America, 84(16): 5818-5822.
Janssens S., Geuten K., Yuan Y-M., Song Y., Küpfer P., and Smets E. (2006). Phylogenetics of Impatiens and Hydrocera (Balsaminaceae) Using Chloroplast atpB-rbcL Spacer Sequences. Systematic Botany, 31(1): 171-180.
Jurado-Rivera J. A., Vogler A. P., Reid C. A. M., Petitpierre E., and Gomez-Zurita J. (2009). DNA barcoding insect–host plant association. Proceedings of the Royal Society B: Biological Sciences, 276: 639-648.
Kan X. Z., Wang S. S., Ding X., and Wang X. Q. (2007). Structural evolution of nrDNA ITS in Pinaceae and its phylogenetic implications. Molecular Phylogenetics and Evolution, 44: 765-777.
Kang Y., Deng Z., Zang R., and Long W. (2017). DNA barcoding analysis and phylogenetic relationships of tree species in tropical cloud forests. Scientific Reports, 7(1): 12564.
Kathriarachchi H., Hoffmann P., Samuel R., Wurdack K. J., and Chase, M. W. (2005). Molecular phylogeneticsofPhyllanthaceae inferredfromfive genes(plastid atpB, matK, 3ndhF, rbcL, andnuclear PHYC). Molecular Phylogenetics and Evolution, 36(1): 34-112.
Kazempour Osaloo S., Kazemi, N. M., Masoumi A. A., and Rastgar P. E. (2006). Phylogenetic status of Oreophysa microphylla (Fabaceae-Galegeae) based on nrDNA (ITS region) and cpDNA (trnL intron/trnL-trnF intergenic spacer) sequences. Rostaniha, 7(226): 177-188.
Kazempour Osaloo S., Maassoumi A. A., and Murakami N. (2003). Molecular systematics of the genus Astragalus L.(Fabaceae): phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacers and chloroplast gene ndhF sequences. Plant Systematics and Evolution, 242(1-4): 1-32.
Kazempour Osaloo S., Maassoumi A. A., and Murakami N. (2005). Molecular systematics of the Old World Astragalus (Fabaceae) as inferred from nrDNA ITS sequence data. Brittonia, 57(4): 367-381.
Keller A., Frster F., Müller T., Dandekar T., Schultz J., and Wolf M. (2010). Including RNA secondary structures improves accu-racy and robustness in reconstruction of phylogenetic trees. Biology Direct, 5: 4.
Kim J. S., Hong J-K., Chase M. W., Fay M. F., and Kim J-H. (2013). Familial relationships of the monocot order Liliales based on a molecular phylogenetic analysis using four plastid loci: matK, rbcL, atpB and atpF-H. Botanical Journal of the Linnean Society, 172(1): 5-21. 
Kim K. J., and Jansen R. K. (1994). Comparisons of phylogenetic hypothesis among different data sets in dwarf dandelions (Krigia): Additional information from internal transcribed spacer sequences of nuclear ribosomal DNA. Plant Systematics and Evolution, 190(3-4): 157-185.
Kim K. J., and Jansen R. K. (1995). NdhF sequence evolution and the major clades in the sunflower Family. Proceedings of the National Academy of Sciences of the United States of America, 92(22): 10379-10383.
Kim S. J., Cho K.S., Yoo K. O., Lim K. B., Hwang Y. J., Chang D. C., and Kim K. S. (2015). Sequence analysis of the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (nrDNA) Chrysanthemum species in Korea. Horticulture, Environment, and Biotechnology. 56: 44-53.
Kjer K. M. (1995). Use of rRNA secondary structure in phylogenetic studies to identify homologous positions: an example of alignment and data presentation from the frogs. Molecular Phylogenetics and Evolution, 4(3): 314–330.
Kron K. (1996). Phylogenetic relationships of Empetraceae, Epacridaceae, and Ericaceae: evidence from nuclear ribosomal 18S sequence data. Annals of Botany, 77: 293-303.
Kuzoff R. K., Sweere J. A., Soltis D. E., Soltis P. S., and Zimmer E. A. (1998). The Phylogenetic Potential of Entire 26S rDNA Sequences in Plants. Molecular Biology and Evolution, 15(3): 251-263.
Lewis C. E., and Doyle J. J. (2002). Phylogenetics of tribe Areceae (Arecaceae) using two low-copy nuclear genes. Plant Systematics and Evolution, 236(1): 1-17.
Li J. H. (2008). Phylogeny of Catalpa (Bignoniaceae) inferred from sequences of chloroplast ndhF and nuclear ribosomal DNA. Journal of Systematics and Evolution, 46(3): 341-348.
Li X., Li Y., Zang M., Li M., and Fang Y. (2018). Complete chloroplast genome sequence and phylogenetic analysis of Quercus acutissima. International Journal of Molecular Sciences, 19(8): 2443.
Li Z., De La Torre A. R., Sterck L., Cánovas F. M., Avila C., Merino I., Cabezas J. A., Cervera M. T., Ingvarsson P. K., and Van de Peer Y. (2017). Single-Copy genes as molecular markers for phylogenomic studies in seed plants. Genome Biology and Evolution, 9(4): 1130-1147.
Linder C. R., Goertzen L. R., Heuvel B. V., Francisco-Ortega J., and Jansen R. K. (2000). The complete external transcribed spacer of 18S-26S rDNA: amplification and phylogenetic utility at low taxonomic levels in Asteraceae and closely allied families. Molecular Phylogenetics and Evolution, 14(2): 285-303.
Little D. P., and Stevenson D. W. (2007). A comparison of algorithms for the identification of specimens using DNA barcodes: Examples from gymnosperms. Cladistics, 23: 1-21.
Liu Q., Brubaker C. L., Green A. G., Marshall D. R., Sharp P. J., and Singh S. P. (2001). Evolution of the FAD2-1 fatty acid desaturase 5′ UTR intron and the molecular systematics of Gossypium (Malvaceae). American Journal of Botany, 88(1): 92-102.
Mäder G., Zamberlan P. M., Fagundes N. J. R., Magnus T., Salzano F. M., Bonatto S. L., and Freitas L. B. (2010). The use and limits of ITS data in the analysis of intraspecific variation in Passiflora L. (Passifloraceae). Genetics and molecular biology, 33: 99-108.
Maggini F., Frediani M., and Gelati M. T. (2000). Nucleotide sequence of the internal transcribed spacers of ribosomal DNA in Picea abies Karst. DNA Sequence, 11(1-2): 87-89.
Maia V. H., Gitzendanner M. A., Soltis P. S., Wong G. K- S., and Soltis D. E. (2014). Angiosperm Phylogeny Based on 18S/26S rDNA Sequence Data: Constructing a Large Data Set Using Next-Generation Sequence Data. International Journal of Plant Sciences, 175(6): 613-650.
Martin G., Baurens F. C., Cardi C., Aury J. M., and D‘Hont A. (2013). The complete chloroplast genome of banana (Musa acuminata, Zingiberales): insight into plastid monocotyledon evolution. PLOS One, 8(2): e67350.
Murphy B. P., and Tranel P. J. (2018). Identification and validation of Amaranthus species-specific SNPs within the ITS region: applications in quantitative species identification. Crop Science, 58: 304-311.
Nei M., and Kumar S. (2000). Molecular Evolution and Phylogenetics, New York: Oxford University Press.
Neyland R. (2001). A phylogeny inferred from large ribosomal subunit (26S) rDNA sequences suggests that Cuscuta is a derived member of Convolvulaceae. Brittonia, 53(1): 108-115.
Nickrent D. L., and Franchina C. R. (1990). Phylogenetic relationships of the Santalales and relatives. Journal of Molecular Evolution, 31: 294-301.
Nickrent D. L., and Soltis D. E. (1995). A comparison of angiosperm phylogenies from nuclear 18S rDNA and rbcL sequences. Annals of the Missouri Botanical Garden, 82: 208-234.
Palmer J. D. (1991). Plastid chromosomes: Structure and evolution. In Bogorad L., and Vasil L. K. (Eds.), Molecular biology of plastids, San Diego, CA, USA: Academic Press, 5-53.
Palmer J. D., Adams K. L., Cho Y., Parkinson C. L., Qiu Y-L., and Song K. (2000). Dynamic evolution of plant mithochondrial genomes: mobile genes and introns and highly variable mutation rates. Proceedings of the National Academy of Sciences of the United States of America, 97(13): 6960-6966.
Pang X., Liu C., Shi L., Liu R., Liang D., Li H., Cherny S. S., and Chen S. (2012) Utility of thetrnH–psbA intergenic spacer region and its combinations as plant DNA barcodes: A meta-analysis. PLoS One, 7(11): e48833.
Peterson P. M., Romaschenko K., and Johnson G. A. (2010). classification of the Chloridoideae (Poaceae) based on multi-gene phylogenetic trees. Molecular Phylogenetics and Evolution, 55(2): 580-598.
Philippe H., Brinkmann H., Lavrov D., Littlewood DTJ., Manuel M, WÖrheide G., and Baurain D. (2011). Resolving Difficult Phylogenetic Questions: Why More Sequences Are Not Enough. PLoS Biology, 9(3): e1000602. doi:10.1371/journal.pbio.1000602.
Queiroz C. S., Batista F. R. C., and Oliveira L. O. (2011). Evolution of the 5.8S nrDNA gene and internal transcribed spacers in Carapichea ipecacuanha (Rubiaceae) within a phylogeographic context. Molecular Phylogenetics and Evolution, 59: 293-302.
Raubeson L. A., and Jansen R. K. (2005). Chloroplast genomes of plants. In Henry R. J. (Ed.), Plant diversity and evolution: genotypic and phenotypic variation in higher plants, Walllingford: CABI, 45-68.
Rieseberg L. H., and Wendel J. F. (1993). Introgression and its consequences in plants. In Harrison R. (Ed.), Hybrid Zones and the Evolutionary Process, Oxford: Oxford University Press, 70-109.
Rieseberg L. H., Baird S. J. E., and Gardner K. A. (2000). Hybridization, introgression, and linkage evolution. Plant Molecular Biology, 42(1): 205-224.
Rogers S. O., and Bendich A. J. (1987). Ribosomal RNA genes in plants: variability in copy number and in intergenic spacer. Plant Molecular Biology, 9(5): 509-520.
Sang T. (2002). Utility of Low-Copy Nuclear Gene Sequences in Plant Phylogenetics. Critical Reviews in Biochemistry and Molecular Biology, 37(3): 121-147.
Sang T., Donoghue M. J., and Zhang D. (1997). Evolution of alcohol dehydrogenase genes in peonies (Paeonia): phylogenetic relationships of putative nonhybrid species. Molecular Biology and Evolution, 14(10): 994-1007.
Sanjur O. I., Piperno D. R., Andres T. C., and Wessel-Beaver L. (2002). Phylogenetic relationships among domesticated and wild species of cucurbita (Cucurbitaceae) inferred from a mithochondrial gene: implications for crop plant evolution and areas of origin. Proceedings of the National Academy of Sciences of the United States of America, 99(1): 535-540.
Schaal B. A., and Olsen K. M. (2000). Gene genealogies and population variation in plants. Proceedings of the National Academy of Sciences of the United States of America, 97(13): 7024-7029.
Senchina D. S., Alvarez I., Cronn R. C., Liu B., Rong J., Noyes R. D., Paterson A. H., Wing R. A., Wilkins T. A., and Wende J. F. (2003). Rate variation among nuclear genes and the age of polyploidy in Gossypium. Molecular Biology and Evolution, 20: 633-643.
Silva G. A. R., Jojima C. L., Moraes E. M., Antonelli A., Manfrin M. H., and Franco F. F. (2016). Intra and interspecific sequence variation in closely related species of Cereus (Cactaceae). Biochemical Systematics and Ecology, 65: 137-142.
Small R. L., Cronn R. C., and Wendel J. F. (2004). Use of nuclear genes for phylogeny reconstruction in plants. Australian Systematic Botany, 17: 145-170.
Small R. L., Ryburn J. A., Cronn R. C., Seelanan T., and Wendel J. F. (1998). The tortoise and the hare: choosing between noncoding plastome and nuclear Adh sequences for phylogenetic reconstruction in a recently diverged plant group. American Journal of Botany, 85(9): 1301-1315.
Soltis D. E., Albert V. A., Leebens-Mack J., Bell C. D., Paterson A. H., Zheng C., Sankoff D., Depamphilis C. W., Wall P. K., and Soltis P. S. (2009). Polyploidy and angiosperm diversification. American Journal of Botany, 96(1): 336-348.
Soltis D. E., Soltis P. S., and Doyle J. J. (1998). Molecular Systematics of Plants II: DNA Sequencing, Boston: Kluwer Aca- demic Publishers.
Soltis D. E., Soltis P. S., Nickrent D. L., Johnson L. A., Hahn W. J., Hoot S. B., Sweere J. A., Kuzoff R. K., Kron K. A., Chase M., Swensen S. M., Zimmer E., Shaw S. M., Gillespie L. J., Kress W. J., and Sytsma K. (1997). Angiosperm phylogeny inferred from 18S ribosomal DNA sequences. Annals of the Missouri Botanical Garden, 84: 1-49.
Soltis P. S., Soltis D. E., Wolf P. G., Nickrent D. L., Chaw S. M., and Chapman R. L. (1999). The phylogeny of land plants inferred from 18S rDNA sequences: pushing the limits of rDNA signal? Molecular Biology and Evolution, 16(12):84-1774.
Stefanovic S., Jager M., Deutsch J., Broutin J., and Masselot M. (1998). Phylogenetic relationships of conifers inferred from partial 28S rRNA gene sequences. American Journal of Botany, 85: 688-697.
Sun J., Shi S., Li J., Yu J., Wang L., Yang X., Guo L., and Zhou S. (2018). Phylogeny of Maleae (Rosaceae) Based on Multiple Chloroplast Regions: Implications to Genera Circumscription. BioMed Research International, 2018: 1-10.
Terra V., Garcia F. C. P., Queiroz L. P. de, Bank M., and Miller J. T. (2017). Phylogenetic Relationships in Senegalia (Leguminosae-Mimosoideae) Emphasizing the South American Lineages. Systematic Botany, 42(3): 458-464.
Toluei A. (2018). The molecular phylogeny of Moltkia Lehm. and Anchusa L., two genera of Boraginaceae using cpDNA trnL-F and nrDNA ITS regions sequence data from Iran. [M.Sc.]. University of Kashan. Kashan. Iran.
Toluei Z., Atri M., Ranjbar M., and Wink M. (2012). Morphological, genetical and ecogeographical characterization of long-winged species of Onobrychis sect. Onobrychis (Fabaceae) in Iran. Iranian Journal of Botany, 18(1): 31-41.
Toluei Z., Atri M., Ranjbar M., and Wink M. (2013a). Iranian Onobrychis carduchorum (Fabaceae) populations: morphology, ecology and phylogeography. Plant Ecology and Evolution, 146(1): 53-67.
Toluei Z., Ranjbar M., Wink M., and Atri M. (2013b). Molecular characterization of Onobrychis altissima (Fabaceae) populations from Iran with the description of O. chaldoranensis, sp. nova. Annales Botanici Fennici, 50: 249-257.
Toluei Z., Ranjbar M., Wink M., and Atri M. (2013c). Molecular phylogeny and ecogeography of Onobrychis viciifolia Scop. (Fabaceae) based on nrDNA ITS sequences and genomic ISSR fingerprinting. Feddes Repertorium, 123(3): 193-207.
Vandamme A. M. (2009). Basic concepts of molecular evolution. In Lemey P., Salemi M. and Vandamme A. M. (Eds.), The phylogenetic Handbook, Cambridge: Cambridge University press, 249-266. 
Wang B., Zhang Y., Wei P., Sun M., Ma X., and Zhu X. (2014). Identification of nuclear low-copy genes and their phylogenetic utility in rosids. Genome, 57(10): 547-554.
Wicke S., Schneeweiss G. M., Müller K. F. and Quandt D. (2011). The evolution of the plastid chromosome in land plants: Gene content, gene order, gene function. Plant Molecular Biology, 76(3-5): 273-297.
Wilson C. A. (2009). Phylogenetic relationships among the recognized series in Iris Section Limniris. Systematic Botany, 34(2): 277-284.
Worning P., Jensen L. J., Nelson K. E., Brunak S., and Ussery D. W. (2000). Structural analysis of DNA sequence: evidence for lateral gene transfer in Thermotoga maritima. Nucleic Acids Research, 28: 706-709.
Yli-Mattila T., Paavanen-Huhtala S., Fenton B., and Tuovinen T. (2000). Species and strain identification of the predatory mite Euseius finlandicus by RAPD-PCR and ITS sequences. Experimental and Applied Acarology, 24(10-11): 863-880.
Zhang N., Zeng L., Shan H., and Ma H. (2012). Highly conserved low-copy nuclear genes as effective markers for phylogenetic analyses in angiosperms. New Phytologist, 195(4): 923-937.
Zhou B., Tu T., Kong F., Wen J., and Xu X. (2018). Revised phylogeny and historical biogeography of the cosmopolitan aquatic plant genus Typha (Typhaceae). Scientific reports, 8: 8813.
Zimmer E. A., and Wen J. (2015). Using nuclear gene data for plant phylogenetics: Progress and prospects II. Next-gen approaches. Journal of Systematics and Evolution, 53(5): 371-379.