Identification and development of microsatellite (SSRs) makers of Exbucklandia (HAMAMELIDACEAE) by high-throughput sequencing
Hamamelidaceae (Saxifragales, previously Rosales) comprises approximately six subfamily, 30 genera and 140 species, most of which are Tertiary relicts. Exbucklandia is the only genus of the subfamily Exbucklandioideae, Hamelidaceae, containing only 2–4 species. Of them, the species E. longipetala H. T. Chang is endemic to China and listed as endangered in The Biodiversity Red List of China: Higher Plant, yet some taxonomists put forward that E. longipetala should be merged into E. tonkinensis (Lecomte) H. T. Chang. Currently, there was nearly no phylogeographic studies on this genus possibly due to the deficiency of efficient molecular markers. In this study, we sequenced the genome of E. tonkinensis based on high throughput sequencing technology, and obtained approximately 6 G raw data, which was further de novo assembled into 303,481 contigs. Based on them, 15,326 SSRs were identified from 13,596 contigs, and primers were successfully designed for 10,660 SSRs. A total of 139 paired primers were synthesized, 106 of them were successfully amplified in six Exbucklandia individuals with expected PCR product size, and 24 demonstrated to be polymorphic among three Exbucklandia populations. Accordingly, the expected and observed heterozygosity were between 0.097–0.717 and 0.098–0.583. Based on these efforts, future researches on genetic diversity and population structure of Exbucklandia can be performed to understand its phylogenetic origins and phylogeographic pattern.
KeywordsExbucklandia Genome sequencing Microsatellite marker Primers development Conservation genetics
We are grateful for collection’s help from Zhao Wanyi, Liu Zhongcheng, the Guangdong Heishiding Natural Reserv and the Limushan National Nature Reserve, Hainan. This work was supported by the National Natural Science Foundation of China (31800175, 31670189, 31570195), Natural Science Foundation of Guangdong Province (2018A0303130109), the Special Program for Science and Technology Basic Research of the Ministry of Science and Technology of China (2013FY111500), the Fourth National Survey on Chinese Material Medical Resources Program for State Administration of Traditional Chinese Medicine of China (2017-152-003) and the Chang Hungta Science Foundation of Sun Yat-sen University.
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Conflict of interest
The author declares there is no conflict of interests.
- 2.Lu A, Zhang ZY (1995) Hamamelidaceae: geographic distribution, fossil history and origin. Acta Phytotaxon Sin 33(4):313–339Google Scholar
- 8.Zhang ZY, Zhang HD (Chang HT), Peter KE (2003) Flora of China. Science Press, Beijing & Missouri Botanical Gardens, St. Louis. Missouri 9: 23–24Google Scholar
- 9.Ministry of Ecology and Environment of the People’s Republic of China, Chinese Academy of Sciences (2013) The biodiversity red list of china: higher plant. http://www.mep.gov.cn/gkml/hbb/bgg/201309/t20130912_260061.htm. Accessed 2 Sept 2013
- 10.Vink W (1957) Flora Malesiana ser. I. The National Herbarium of the Netherlands 5:363–379Google Scholar
- 11.Lakhanpal RN (1958) The Rujada flora of west central Oregon. University of California Publications in Geological Sciences 35:1–66Google Scholar
- 13.Smiley C (1985) Late Cenozoic history of the Pacific Northwest: interdisciplinary studies on the Clarkia fossil bed. Pacific Division of the American Association for the Advancement of Science 95–112Google Scholar
- 15.Li ZM, Yang XS (1978) Fine tree species: Exbuaklandia. Guangxi For. Sci. 4:13–14Google Scholar
- 17.Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem. Bull. 19:11–15Google Scholar
- 20.Rozen S, Skaletsky H (1999) Primer3 on the WWW for general users and for biologist programmers. In Bioinformatics methods and protocols. Humana Press, Totowa, p. 365Google Scholar