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Parasitic-Relationship-Of-Cistanche-Deserticola-And-Host-Pl 2019 Chinese-Her.Pdf Chinese Herbal Medicines 11 (2019) 267–274 Contents lists available at ScienceDirect Chinese Herbal Medicines journal homepage: www.elsevier.com/locate/chmed Original Article Parasitic relationship of Cistanche deserticola and host-plant Haloxylon ammodendron based on genetic variation of host ∗ Liang Shen a,b, Rong Xu a, , Sai Liu a, Chang-qing Xu a, Fang Peng a, Xiao-jin Li c, ∗ Guo-qiang Zhu c, Cai-xiang Xie a, Jun Zhu c, Tong-ning Liu d, Jun Chen a, a Institute of Medicinal Plant Development, Chinese Academy of Medical Science & Peking Union Medical College, Beijing 100193, China b Beijing Museum of Natural History, Beijing Academy of Science and Technology, Beijing 10 0 050, China c Xinjiang Institutes of Traditional Chinese Medica and National Medica, Urumqi 830 0 02, China d Ningxia Yongning Plantation of Herba Cistanche, Yinchuan 750100, China a r t i c l e i n f o a b s t r a c t Article history: Objective: Cistanche deserticola is a famous and endangered medicinal plant that is parasitic upon Haloxy- Received 5 May 2018 lon ammodendron with rather low parasitic rates . It is important to find high affinity germplasms for Revised 13 January 2019 increasing the survival of C. deserticola . However, little is known in genetic variation and high affinity Accepted 16 April 2019 populations of H. ammodendron in China. Available online 6 June 2019 Methods: In this study, 98 accessions of H. ammodendron seeds were collected from five regions covering Keywords: almost the entire natural distribution of H. ammodendron in China. Their genetic variations were analyzed Cistanche deserticola Y. C. Ma using AFLP and ITS by the maximum parsimony method, and a dendrogram was constructed using the genetic structures unweighted pair-group method with arithmetic average (UPGMA). The parasitic rates of C. deserticola on Haloxylon ammodendron (C. A. Mey.) Bunge different accessions of H. ammodendron were calculated in the field experiment. parasitic rates Results: Both AFLP and ITS methods consistently revealed that there was a high level of genetic di- versity in the natural populations of H. ammodendron . Hierarchical population structure analysis uncov- ered a clear pattern that all populations were grouped into three main clusters, and eight populations from eastern region were genetically clustered together. These regions were significantly differentiated ( P < 0.05), 13.10% of variation occurred among populations, and 86.90% within populations was revealed by analysis of molecular variance (AMOVA). The populations of Inner Mongolia had the highest parasitic rates followed by Ganjiahu Reserve and Yongning Plantation for the top three, which were not completely related to the genetic variation. Conclusion: Genetic characteristics of H. ammodendron in China were clarified and the order of affinity of different populations was given, which were primers for discovering high affinity germplasms. ©2019 Tianjin Press of Chinese Herbal Medicines. Published by Elsevier B.V. All rights reserved. 1. Introduction Genetic variation is one of the premiers for conserving host and parasite, with view to screening out of high affinity germplasms Cistanche deserticola Y. C. Ma is a rare and famous medicinal ( Bay et al., 2017; Cortanˇ & Tubi c,´ 2017 ). Several molecular markers plant which mainly parasitizes on roots of Haloxylon ammoden- including isozyme, ISSR, RAPD and AFLP have been used to study dron (C. A. Mey.) Bunge, a xerophytic desert shrub (Amaranthaceae) genetic variation of H. ammodendron populations ( Sheng, Zheng, ( Akhani, Edwards, & Roalson, 2007; Toghraie, 2012 ). Due to a low & Pei, 2005; Zhang, Dong, Wei, & Hu, 2006; Wang et al., 2009 ; parasitic rate upon host H. ammodendron, C. deserticola is an en- Shen et al., 2014 , 2015 ) . However, population samples in those dangered species that are deteriorated by land reclamation, over- studies were not enough for revealing genetic spectrum and par- cutting, and overstocking ( Wang et al., 2015; Xu et al., 2009 ). In asitic rates in the unclear field. It is necessary to collect more pop- China, H. ammodendron has been reduced from 110 0 0 0 km 2 in the ulations to represent entire H. ammodendron germplasms in China 1960s to 65 300 km 2 ( Guo et al., 2005 ) . It is important to find the and investigate the parasitic rates of different populations. host-plant germplasms with high parasitic rates for the survival of In recent years, AFLP markers have been used to detect the ge- C. deserticola . netic variation and structure of many species ( Shen et al., 2014; Zhao, Yin, Xu, & Wang, 2012, Głebocka ˛ & Pogorzelec, 2017 ). The AFLP technique provided genetic information on multiple loci in a ∗ Corresponding authors. E-mail addresses: [email protected] (R. Xu), [email protected] (J. Chen). single assay without prior sequence knowledge at a relatively low https://doi.org/10.1016/j.chmed.2019.04.006 1674-6384/© 2019 Tianjin Press of Chinese Herbal Medicines. Published by Elsevier B.V. All rights reserved. 268 L. Shen, R. Xu and S. Liu et al. / Chinese Herbal Medicines 11 (2019) 267–274 cost with the requirement of small quantities of plant materials ering almost the entire distribution of the species in China. The ( Joyce et al., 2014 ). Another DNA sequence, from the internal tran- results will contribute to the accessions of H. ammodendron with a scribed spacer (ITS) region of the nuclear genome, is commonly high affinity with C. deserticola. applied in species identification, genetic variation and phylogenetic evolutionary studies ( Chen et al., 2010 ). ITS sequences are prone 2. Materials and methods to neutral mutations without strong constraints, leading to a rela- tively fast evolutionary rate and making the region appropriate for 2.1. Plant materials phylogenetic and evolutionary studies ( Hebert, Cywinska, & Ball, 2003 ). Assessing the evolutionary phylogenetic of H. ammodendron A total of 98 accessions of H. ammodendron were collected using AFLP jointly with ITS sequence can provide a valuable ge- from 14 populations from five regions including Xinjiang, Gansu, netic complementary information about host-plant identification, Ningxia, Qinghai, and Inner Mongolia, located between longitudes genetic variation and structure of a population. 82 °E to 106 °E and latitudes 36 °N to 44 °N, approximately covering In this study, the genetic variation and parasitic relationship of the main area of the species in China ( Fig. 1 , Table 1 ). Reserves H. ammodendron and C. deserticola were analyzed, and 14 popu- of Ganjiahu (Tuoli, Huyang and Guanfengtai), Tsaidam (Bolanggou, lations of H. ammodendron from five regions including Xinjiang, Huaitoutala and Zhaohe) and Inner Mongolia (Urat Reserve, Or- Gansu, Ningxia, Qinghai, and Inner Mongolia were collected, cov- dos Reserve and Alxa Reserve) were national nature reserves, and Fig. 1. Geographical distribution of five growing regions [XG (XGT, XGH and XGG); QT (QTB, QTH and QTZ); GL (GLG, GLH and GLS); IM (IMUB, IMOK and IMAJ); NY (NYG and NYY)] of H. ammodendron sampled in this study. Populations were represented by black dots and their locations were given in Table 1 . Map was plotted using software ArcGIS 10.2 (ESRI, Redland, CA. URL http://www.esri.com/ ). Table 1 Samples of Haloxylon ammodendron used in this study. No Region codes Population codes Location of populations Longitude / °E Latitude / °N Altitude / m Samples No. 1 XG XGT Ganjiahu Reserve Tuoli Xinjiang 82.59 44.76 277.40 7 2 XG XGH Ganjiahu Reserve Huyang Xinjiang 83.43 44.91 254.60 7 3 XG XGG Ganjiahu Reserve Guanfengtai 83.41 44.88 249.50 7 4 QT QTB Tsaidam Reserve Bolanggou 97.74 36.60 2902.40 7 5 QT QTH Tsaidam Reserve Huaitoutala 96.55 37.09 3123.60 7 6 QT QTZ Tsaidam Reserve Zhaohe 97.67 36.77 2960.30 7 7 GL GLG Liangucheng Reserve Qinfeng 102.95 38.61 1366.30 7 8 GL GLH Liangucheng Reserve Huaer 102.45 38.83 1354.90 7 9 GL GLS Liangucheng Reserve Xuebai 102.99 38.74 1379.00 7 10 IM IMUB Inner Mongolia 105.80 41.23 1068.60 7 Urat Reserve 11 IM IMOK Inner Mongolia 107.61 40.37 1084.00 7 Ordos Reserve 12 IM IMAJ Inner Mongolia 105.77 39.61 1048.00 7 Alxa Reserve 13 NY NYG Yongning Plantation Guangxia 106.08 38.24 1124.20 7 14 NY NYY Yongning Plantation Yuquan 106.02 38.22 1130.90 7 Note: XG, Xinjiang Ganjiahu; QT, Qinghai Tsaidam; GL, Gansu Liangucheng; IM, Inner Mongolia; NY, Ningxia Yongning. L. Shen, R. Xu and S. Liu et al. / Chinese Herbal Medicines 11 (2019) 267–274 269 Liangucheng reserve (Qinfeng, Huaeryuan and Xuebai) and Yongn- cessions was based on Jaccard’s genetic distance matrix of acces- ing plantation (Guangxia and Yuquan) were man-made in the 1960 sion; Data were subjected to genetic analysis in GenAlEx package and 1998, respectively. Seeds and seedlings of H. ammodendron at ( Peakall & Smouse, 2012 ). Yongning plantation were mainly portion of Inner Mongolia. The A Bayesian approach (Markov Chain Monte Carlo Algorithm) accessions of H. ammodendron were identified by Professor Jun was applied to detect the underlying population genetic structure Chen, and the voucher specimens (H2015-1) were deposited at the among a set of accessions ( Wolf, Anselin, & Arribas, 2018 ). The National Medicinal Plant Seed Resource Library of the Institute of most likely number of clusters was estimated according to the Medicinal Plant Development, Chinese Academy of Medical Sci- model value ( K ) based on the second order rate of change, fol- ences (Beijing), China. lowing the method described by Sorkheh, Dehkordi, Ercisli, Hege- dus, and Halász (2017) . Five STRUCTURE runs were performed 2.2. DNA extraction and PCR for each K , ranging from 1 to 15, independently, with a burn- in length of 10 0 0 0 followed by 100 000 runs at each K , and a Seven accessions of each population were randomly selected higher run from the five runs was chosen.
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