Advertisement

Trees

, Volume 32, Issue 6, pp 1559–1571 | Cite as

Development of EST-SSR markers in Larix principis-rupprechtii Mayr and evaluation of their polymorphism and cross-species amplification

  • Mingliang Dong
  • Zewei Wang
  • Qingwei He
  • Jian Zhao
  • Zhirong Fan
  • Jinfeng ZhangEmail author
Original Article

Abstract

Key message

We developed and validated a new set of polymorphic EST-SSR markers across Larix species and evaluated genetic diversity in a clonal seed orchard of Larix principis-rupprechtii Mayr.

Abstract

Prince Rupprecht’s larch (Larix principis-rupprechtii Mayr) is an important deciduous conifer species that has been widely planted in North China due to its major ecological and commercial value. However, the paucity of genomic data and robust molecular markers has hampered genetic and genomic studies. Here, transcriptome sequencing of L. principis-rupprechtii callus was performed using the Illumina platform. By mining 43,753 assembled unigenes, 1418 expressed sequence tag-simple sequence repeats (EST-SSRs) derived from 1300 unigenes were identified. A total of 1065 primer pairs were designed and 240 of these selected at random for validation among 24 L. principis-rupprechtii individuals. Of these, 52 primer pairs were scored as polymorphic, and 20 polymorphic EST-SSR markers were further selected to genotype 66 clones deployed in a clonal seed orchard of L. principis-rupprechtii; these exhibited a moderate level of genetic diversity, as reflected by the mean values of the number of alleles (Na = 3.85) and polymorphism information content (PIC = 0.424). Additionally, all of the 20 EST-SSR markers could amplify clear and stable bands across three related Larix species. A neighbor-joining (NJ) clustering tree uniquely distinguished 66 clones and distributed these into three main clusters, which was further validated by principal coordinate analysis (PCoA). The developed EST-SSR markers will serve as valuable tools for future genetics and breeding research in larch species. The evaluation of genetic diversity among 66 clones will provide important information for efficient management and utilization of genetic material in L. principis-rupprechtii breeding programs.

Keywords

EST-SSR markers Genetic diversity Larix principis-rupprechtii Transcriptome Transferability 

Notes

Acknowledgements

This paper was supported by National Key R&D Program of China (2017YFD0600404-1), Medium and Long Scientific Research Project for Young Teachers in Beijing Forestry University (2015ZCQ-SW-02), the Project of National Natural Science Foundation of China (31370658), “948” Project of China (2014-4-59), and major science and technology special project of Xuchang, Henan Province, China (20170112006). We are grateful to the Fine Variety Base of Longtoushan (Weichang County, China) and Jingle County Forestry Bureau (Jingle County, China) for their assistance in collecting samples.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Data archiving statement

The RNA sequencing raw data have been submitted to the NCBI Short Read Archive (SRA) with the accession number SRP127010.

Supplementary material

468_2018_1733_MOESM1_ESM.xlsx (35 kb)
Supplementary material 1 (XLSX 34 KB)

References

  1. Aggarwal RK, Hendre PS, Varshney RK, Bhat PR, Krishnakumar V, Singh L (2007) Identification, characterization and utilization of EST-derived genic microsatellite markers for genome analyses of coffee and related species. Theor Appl Genet 114:359–372CrossRefGoogle Scholar
  2. Azevedo H, Lino-Neto T, Rui MT (2003) An improved method for high-quality RNA isolation from needles of adult maritime pine trees. Plant Mol Biol Rep 21:333–338CrossRefGoogle Scholar
  3. Botstein D, White RL, Skolnick M, Davis RW (1980) Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am J Hum Genet 32:314–331PubMedPubMedCentralGoogle Scholar
  4. Canales J, Bautista R, Label P et al (2014) De novo assembly of maritime pine transcriptome: implications for forest breeding and biotechnology. Plant Biotechnol J 12:286–299CrossRefGoogle Scholar
  5. Chen C, Liewlaksaneeyanawin C, Funda T, Kenawy A, Newton CH, El-kassaby YA (2009) Development and characterization of microsatellite loci in western larch (Larix occidentalis Nutt.). Mol Ecol Resour 9:843–845CrossRefGoogle Scholar
  6. Chen XB, Xie YH, Sun XM (2015) Development and characterization of polymorphic genic-SSR markers in Larix kaempferi. Molecules 20:6060–6067CrossRefGoogle Scholar
  7. Cheng YL, Yang Y, Wang ZY, Qi BY, Yin YL, Li HG (2015) Development and characterization of EST-SSR markers in Taxodium ‘zhongshansa’. Plant Mol Biol Rep 33:1–11CrossRefGoogle Scholar
  8. Colburn BC, Mehlenbacher SA, Sathuvalli VR (2017) Development and mapping of microsatellite markers from transcriptome sequences of European hazelnut (Corylus avellana L.) and use for germplasm characterization. Mol Breed 37:16CrossRefGoogle Scholar
  9. Di XY, Li XN, Wang QX, Wang MB (2014) Genetic diversity of natural populations of Larix principis-rupprechtii, in Shanxi Province, China. Biochem Syst Ecol 54:71–77CrossRefGoogle Scholar
  10. Dong ML, Gao JY, Sun WT, Fan YM, Yuan DC, Zhang JF (2016) Genetic diversity and population structure of Larix principis rupprechtii Mayr in Beijing. J Plant Genet Resour 17:616–624 (in Chinese) Google Scholar
  11. Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh leaf tissue. Phytochem Bull 19:11–15Google Scholar
  12. Du J, Zhang Z, Zhang HG, Tang JH (2017) EST-SSR marker development and transcriptome sequencing analysis of different tissues of Korean pine (Pinus Koraiensis Sieb. et Zucc.). Biotechnol Biotec Eq 31:679–689Google Scholar
  13. Durand J, Bodénès C, Chancerel E et al (2010) A fast and cost-effective approach to develop and map EST-SSR markers: oak as a case study. BMC Genom 11:570CrossRefGoogle Scholar
  14. Dutta S, Kumawat G, Singh BP et al (2011) Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]. BMC Plant Biol 11:17CrossRefGoogle Scholar
  15. Fan YM, Zhang DR, Yu DD et al (2014) Genetic diversity and population structure of Larix principis- rupprechtii Mayr in Hebei Province. J Plant Genet Resour 15:465–471 (in Chinese) Google Scholar
  16. Funda T, Lstibůrek M, Lachout P, Klápště J, El-Kassaby YA (2009) Optimization of combined genetic gain and diversity for collection and deployment of seed orchard crops. Tree Genet Genome 5:583–593CrossRefGoogle Scholar
  17. Garg R, Patel RK, Tyagi AK, Jain M (2011) De novo assembly of chickpea transcriptome using short reads for gene discovery and marker identification. DNA Res 18:53–63CrossRefGoogle Scholar
  18. Grabherr MG, Haas BJ, Yassour M et al (2011) Trinity: reconstructing a full-length transcriptome without a genome from RNA-Seq data. Nat Biotechnol 29:644CrossRefGoogle Scholar
  19. Hao W, Wang SJ, Liu HJ, Zhou BR, Wang XW, Jiang TB (2015) Development of SSR markers and genetic diversity in white birch (Betula platyphylla). PLoS One 10:e0129758CrossRefGoogle Scholar
  20. Hu XG, Liu H, Jin YQ et al (2016) De novo transcriptome assembly and characterization for the widespread and stress-tolerant conifer Platycladus orientalis. PLoS One 11:e0148985CrossRefGoogle Scholar
  21. Isoda K, Watanabe A (2006) Isolation and characterization of microsatellite loci from Larix kaempferi. Mol Ecol Resour 6:664–666CrossRefGoogle Scholar
  22. Jia GX, Shen XH (2001) Study on pollination biology of Larix principis-rupprechtii Mayr. Scientia Silvae Sinicae 37:40–45 (in Chinese) Google Scholar
  23. Jia GX, Yang JM, Shen XH (2003) Variation in fruiting ability of interspecific crossing and mechanism of self-depression in Larix. Scientia Silvae Sinicae 39:62–68 (in Chinese) Google Scholar
  24. Jia XP, Deng YM, Sun XB, Liang LJ, Su JL (2016) De novo assembly of the transcriptome of Neottopteris nidus, using Illumina paired-end sequencing and development of EST-SSR markers. Mol Breed 36:1–12CrossRefGoogle Scholar
  25. Kalinowski ST, Taper ML, Marshall TC (2007) Revising how the computer program cervus accommodates genotyping error increases success in paternity assignment. Mol Ecol 16:1099–1106CrossRefGoogle Scholar
  26. Khasa PD, Newton CH, Rahman MH, Jaquish B, Dancik BP (2000) Isolation, characterization, and inheritance of microsatellite loci in alpine larch and western larch. Génome 43:439–448CrossRefGoogle Scholar
  27. Kumar S, Stecher G, Tamura K (2016) MEGA7: molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol Biol Evol 33:1870–1874CrossRefGoogle Scholar
  28. Lai HL, Wang ZR (1997) Comparison of genetic structure between parents and progeny from a masson pine seed orchard and a plantation nearby. For Res 10:490–494 (in Chinese) Google Scholar
  29. Lelu-Walter MA, Pâques LE (2009) Simplified and improved somatic embryogenesis of hybrid larches (Larix × eurolepis and Larix × marschlinsii). Perspectives for breeding. Ann For Sci 66:104CrossRefGoogle Scholar
  30. Li Y, Zhang CX (2000) Genetic diversity within a breeding system of Pinus tabulaeformis. J Beijing For Univ 22:12–19 (in Chinese)Google Scholar
  31. Li DJ, Deng Z, Qin B, Liu XH, Men ZH (2012) De novo assembly and characterization of bark transcriptome using Illumina sequencing and development of EST-SSR markers in rubber tree (Hevea brasiliensis Muell. Arg.). BMC Genom 13:192CrossRefGoogle Scholar
  32. Liu K, Muse SV (2005) PowerMarker: an integrated analysis environment for genetic marker analysis. Bioinformatics 21:2128–2129CrossRefGoogle Scholar
  33. Mariotti R, Cultrera NGM, Mousavi S et al (2016) Development, evaluation, and validation of new EST-SSR markers in olive (Olea europaea L.). Tree Genet Genome 12:120CrossRefGoogle Scholar
  34. Metzgar D, Bytof J, Wills C (2000) Selection against frameshift mutations limits microsatellite expansion in coding DNA. Genome Res 10:72–80PubMedPubMedCentralGoogle Scholar
  35. Millar MA, Byrne M, Nuberg I, Sedgley M (2008) High outcrossing and random pollen dispersal in a planted stand of Acacia saligna subsp. saligna revealed by paternity analysis using microsatellites. Tree Genet Genome 4:367–377CrossRefGoogle Scholar
  36. Nei M, Tajima F, Tateno Y (1983) Accuracy of estimated phylogenetic trees from molecular data II. Gene frequency data. J Mol Evol 19:153–170CrossRefGoogle Scholar
  37. Niu SH, Li ZX, Yuan HW, Chen XY, Li Y, Li W (2013) Transcriptome characterisation of Pinus tabuliformis and evolution of genes in the Pinus phylogeny. BMC Genom 14:263CrossRefGoogle Scholar
  38. Parchman TL, Geist KS, Grahnen JA, Benkman CW, Buerkle CA (2010) Transcriptome sequencing in an ecologically important tree species: assembly, annotation, and marker discovery. BMC Genom 11:1–16CrossRefGoogle Scholar
  39. Peakall R, Smouse PE (2006) GENALEX 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Resour 6:288–295CrossRefGoogle Scholar
  40. Pinosio S, González-Martínez SC, Bagnoli F et al (2014) First insights into the transcriptome and development of new genomic tools of a widespread circum-Mediterranean tree species, Pinus halepensis Mill. Mol Ecol Resour 14:846–856CrossRefGoogle Scholar
  41. Poltri SNM, Zelener N, Traverso JR, Gelid P, Hopp HE (2003) Selection of a seed orchard of Eucalyptus dunnii based on genetic diversity criteria calculated using molecular markers. Tree Physiol 23:625–632CrossRefGoogle Scholar
  42. Postolache D, Leonarduzzi C, Piotti A et al (2014) Transcriptome versus genomic microsatellite markers: highly informative multiplexes for genotyping Abies alba Mill. and congeneric species. Plant Mol Biol Rep 32:750–760CrossRefGoogle Scholar
  43. Rozen S, Skaletsky H (2000) Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol 132:365–386PubMedGoogle Scholar
  44. Schuelke M (2000) An economic method for the fluorescent labeling of PCR fragments. Nat Biotechnol 18:233–234CrossRefGoogle Scholar
  45. Sun WT, Yu DD, Dong ML et al (2017) Evaluation of efficiency of controlled pollination based parentage analysis in a Larix gmelinii var. principis-rupprechtii Mayr. seed orchard. PLoS One 12:e0176483CrossRefGoogle Scholar
  46. Thiel T, Michalek W, Varshney RK, Graner A (2003) Exploiting EST databases for the development and characterization of gene-derived SSR-markers in barley (Hordeum vulgare L.). Theor Appl Genet 106:411–422CrossRefGoogle Scholar
  47. Ueno S, Moriguchi Y, Uchiyama K et al (2012) A second generation framework for the analysis of microsatellites in expressed sequence tags and the development of EST-SSR markers for a conifer, cryptomeria japonica. BMC Genom 13:136CrossRefGoogle Scholar
  48. Varshney RK, Graner A, Sorrells ME (2005) Genic microsatellite markers in plants: features and applications. Trends Biotechnol 23:48–55CrossRefGoogle Scholar
  49. Xiang XY, Zhang ZX, Wang ZG, Zhang XP, Wu GL (2015) Transcriptome sequencing and development of EST-SSR markers in Pinus dabeshanensis, an endangered conifer endemic to China. Mol Breed 35:158CrossRefGoogle Scholar
  50. Xu YB, Crouch JH (2008) Marker-assisted selection in plant breeding: from publications to practice. Crop Sci 48:391–407CrossRefGoogle Scholar
  51. Xu CL, Sun XM, Zhang SG (2013) Characteristics of conifer genome and recent advances in conifer sequence resources mining. Chin Bull Bot 48:684–693 (in Chinese) Google Scholar
  52. Yang JM, Shen XH (2002) Individual and family variations in rooting ability of Larix principis-rupprechtii Mayr. J Beijing For Univ 24:8–13 (in Chinese)Google Scholar
  53. Yang X, Sun X, Zhang S (2011) Short note: development of six EST-SSR markers in larch. Silvae Genet 60:161–163CrossRefGoogle Scholar
  54. Yu DD, Yuan DC, Zhang DR et al (2014) Genetic diversity of Larix principis-rupprechtii Mayr. seed orchard among generations. J Plant Genet Resour 15:940–947 (in Chinese) Google Scholar
  55. Zalapa JE, Cuevas H, Zhu H et al (2012) Using next-generation sequencing approaches to isolate simple sequence repeat (SSR) loci in the plant sciences. Am J Bot 99:193–208CrossRefGoogle Scholar
  56. Zhai LL, Xu L, Wang Y et al (2014) Novel and useful genic-SSR markers from de novo transcriptome sequencing of radish (Raphanus sativus L.). Mol Breed 33:611–624CrossRefGoogle Scholar
  57. Zhang XB, Ren JR, Zhang DE (2001) Phenological observations on Larix principis-rupprechtii Mayr. in primary seed orchard. J For Res 12:201–204CrossRefGoogle Scholar
  58. Zhang HJ, Dai JF, Lin Y (2015a) Variation of crossability and seed shape of Larix principis-rupprechtii and Larix kaempferi interspecific hybrids. J Northwest A&F Univ (Nat Sci Ed) 43:36–42 (in Chinese) Google Scholar
  59. Zhang GJ, Sun ZZ, Zhou D et al (2015b) Development and characterization of novel EST-SSRs from Larix gmelinii and their cross-species transferability. Molecules 20:12469–12480CrossRefGoogle Scholar
  60. Zhao J, Wang BB, Wang XQ, Zhang Y, Dong ML, Zhang JF (2015) iTRAQ-based comparative proteomic analysis of embryogenic and non-embryogenic tissues of Prince Rupprecht’s larch (Larix principis-rupprechtii Mayr). Plant Cell Tiss Org 120:655–669CrossRefGoogle Scholar
  61. Zonneveld BJM (2012) Conifer genome sizes of 172 species, covering 64 of 67 genera, range from 8 to 72 picogram. Nord J Bot 30:490–502CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Mingliang Dong
    • 1
    • 2
    • 3
  • Zewei Wang
    • 1
    • 2
    • 3
  • Qingwei He
    • 1
    • 2
    • 3
  • Jian Zhao
    • 1
    • 2
    • 3
  • Zhirong Fan
    • 4
  • Jinfeng Zhang
    • 1
    • 2
    • 3
    Email author
  1. 1.Beijing Advanced Innovation Center for Tree Breeding by Molecular Design, National Engineering Laboratory for Tree Breeding, Key Laboratory of Genetics and Breeding in Forest Trees and Ornamental Plants of Ministry of EducationBeijing Forestry UniversityBeijingChina
  2. 2.Key Laboratory of Forest Trees and Ornamental Plants Biological Engineering of State Forestry AdministrationBeijing Forestry UniversityBeijingChina
  3. 3.College of Biological Sciences and BiotechnologyBeijing Forestry UniversityBeijingChina
  4. 4.Jingle County Forestry BureauJinglePeople’s Republic of China

Personalised recommendations