Genomic insights into salt adaptation in a desert poplar

ARTICLE Received 1 Sep 2013 | Accepted 21 Oct 2013 | Published 21 Nov 2013 | Updated 17 Mar 2014 DOI: 10.1038/ncomms3797 Genomic insights into salt adaptation in a desert poplar Tao Ma1,*, Junyi Wang2,*, Gongke Zhou3,*, Zhen Yue2, Quanjun Hu1, Yan Chen2, Bingbing Liu1, Qiang Qiu1, Zhuo Wang2, Jian Zhang1, Kun Wang1, Dechun Jiang1, Caiyun Gou2, Lili Yu2, Dongliang Zhan2, Ran Zhou1, Wenchun Luo1, Hui Ma1, Yongzhi Yang1, Shengkai Pan2, Dongming Fang2, Yadan Luo2, Xia Wang1, Gaini Wang1, Juan Wang1, Qian Wang1, Xu Lu1, Zhe Chen2, Jinchao Liu2, Yao Lu2, Ye Yin2, Huanming Yang2, Richard J. Abbott4, Yuxia Wu1, Dongshi Wan1, Jia Li1, Tongming Yin5, Martin Lascoux6, Stephen P. DiFazio7, Gerald A. Tuskan8, Jun Wang2,9 & Jianquan Liu1 Despite the high economic and ecological importance of forests, our knowledge of the genomic evolution of trees under salt stress remains very limited. Here we report the genome sequence of the desert poplar, Populus euphratica, which exhibits high tolerance to salt stress. Its genome is very similar and collinear to that of the closely related mesophytic congener, P. trichocarpa. However, we find that several gene families likely to be involved in tolerance to salt stress contain significantly more gene copies within the P. euphratica lineage. Furthermore, genes showing evidence of positive selection are significantly enriched in functional categories related to salt stress. Some of these genes, and others within the same categories, are significantly upregulated under salt stress relative to their expression in another salt-sensitive poplar. Our results provide an important background for understanding tree adaptation to salt stress and facilitating the genetic improvement of cultivated poplars for saline soils. 1 State Key Laboratory of Grassland Agro-Ecosystem, School of Life Sciences, Lanzhou University, Lanzhou 730000, China. 2 BGI-Shenzhen, Shenzhen 518083, China. 3 Key Laboratory of Biofuels and Shandong Provincial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao 266101, China. 4 School of Biology, Mitchell Building, University of St Andrews, St Andrews, Fife KY16 9TH, UK. 5 The Key Lab of Forest Genetics and Gene Engineering, Nanjing Forestry University, Nanjing 210037, China. 6 Department of Ecology and Genetics, Evolutionary Biology Centre, Uppsala University, Norbyvägen, 18D 75326 Uppsala, Sweden. 7 Department of Biology, West Virginia University, Morgantown, West Virginia 26506-6057, USA. 8 BioSciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee 37831, USA. 9 Department of Biology, University of Copenhagen, Copenhagen 1017, Denmark. * These authors contributed equally to this work. Correspondence and requests for materials should be addressed to J.L. (email: ) or to J.W. (email: ). NATURE COMMUNICATIONS | 4:2797 | DOI: 10.1038/ncomms3797 | www.nature.com/naturecommunications & 2013 Macmillan Publishers Limited. All rights reserved. 1 ARTICLE NATURE COMMUNICATIONS | DOI: 10.1038/ncomms3797 orests dominate much of the terrestrial landscape1. However, forest trees rarely occur on saline soils and little is known of the genetic basis of their tolerance to salt stress despite strong demand for their cultivation on highly saline soils in many parts of the world2. Members of the genus Populus are used as a model forest species for diverse studies not only because of their amenability to experimental and genetic manipulation, but also because of their high economic and ecological importance as the most widely cultivated tree throughout the northern hemisphere3,4. More than 30 wild Populus species occur across diverse habitats over a wide geographical range, thereby providing an excellent system for unravelling the genetic bases of adaptive divergence4. Populus euphratica Oliv., which is native to desert regions ranging from western China to North Africa, is characterized by extraordinary adaptation to salt stress5–8. Notably, at high salinity it maintains higher growth and photosynthetic rates than other poplar species9,10 and can survive concentrations of NaCl in nutrient solution up to 450 mM11. In this study, we examine genomic differences between a xeric desert poplar and its mesophytic congener, P. trichocarpa, for which a high-quality reference genome is available12. We further examine gene expression differences following salt stress treatment in a comparison with another salt-sensitive congener, P. tomentosa. Our comparisons highlight the genetic bases of salt tolerance in the desert poplar. F Results Genome sequencing and assembly. Because of the limitations of next-generation sequencing for complex genome assembly13 and the high levels of polymorphism found in this non-domesticated and open-pollinated species (Supplementary Fig. S1), we employed a newly developed fosmid-pooling strategy14 to sequence and assemble the P. euphratica genome (Table 1 and Supplementary Methods). Hierarchical assembly using 67.1 Gb (B112  ) wholegenome shotgun reads (Supplementary Table S1), combined with more than 200  high-quality reads from 66,240 fosmid clones (Supplementary Table S2), yielded a final assembly with a total length of 496.5 Mb (Supplementary Table S3), representing 83.7% of the P. euphratica nucleotide space (Supplementary Tables S4 and S5). The contig N50 of the assembled sequence was 40.4 Kb (longest, 728.4 Kb) and scaffold N50 was 482 Kb (longest, 8.8 Mb; Table 1), which were comparable to those of other plant genome assemblies generated by next-generation sequencing technology (Supplementary Table S6). Sequencing depth distribution showed that over 92.5% of the assembly was covered by more than 20  (Supplementary Figs S2 and S3), ensuring a high single-base accuracy. The heterozygosity level in P. euphratica was B0.5% (Supplementary Tables S7 and S8, and Supplementary Fig. S4), which is almost twice that in P. trichocarpa (0.26%)12. The assembly covered 97.3% of the 516,712 Populus expressed sequence tags (Supplementary Table S9) and 97.7% of the 7 complete fosmids sequenced by Sanger sequencing (Supplementary Table S10 and Supplementary Fig. S5), without any obvious misassembly occurring. The coverage of the core eukaryotic genes was estimated to be 94.35% for the P. euphratica assembly (Supplementary Table S11), which is comparable to the estimate for P. trichocarpa (93.95%). All of these statistics supported that our draft genome sequence has high contiguity, coverage and accuracy, further demonstrating the feasibility of this hierarchical approach for de novo sequencing and assembly of a complex genome with high heterozygosity14. Genome annotation. Using a combination of homology-based searches and de novo annotation, we found that B44% of the P. euphratica genome is composed of repetitive elements (Supplementary Table S12), similar to that of the P. trichocarpa genome (47%; Fig. 1). Long-terminal repeats (LTRs) were th (...truncated)