Construction of molecular ID for Osmanthus fragrans cultivars based on phenotypic traits and single nucleotide polymorphisms (SNPs)

WANG Yihan, LIU Jiaojiao, JIN Peiquan, LI Shuqing, WEI Jianfen, GUO Peng, SHANG Fude

JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2024, Vol. 48 ›› Issue (4) : 12-24.

PDF(5559 KB)
PDF(5559 KB)
JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2024, Vol. 48 ›› Issue (4) : 12-24. DOI: 10.12302/j.issn.1000-2006.202405026

Construction of molecular ID for Osmanthus fragrans cultivars based on phenotypic traits and single nucleotide polymorphisms (SNPs)

Author information +
History +

Abstract

【Objective】This study selected core genomic single-nucleotide polymorphism (SNP) loci to establish a rapid SNP genotyping method on the KASP platform, and to construct molecular IDs for Osmanthus fragrans cultivars. This study provides a theoretical foundation for identifying, tracing and protecting the intellectual property of O. fragrans cultivars.【Method】Field surveys were conducted to investigate key phenotypic characteristics of O. fragrans cultivars. Following two rounds of rigorous screening, we identified a set of core SNP markers capable of completely distinguishing previously sequenced cultivars. Subsequently, we analyzed the polymorphic information content (PIC) and expected heterozygosity (He) of each SNP locus. Using the genome sequences of ‘Rixianggui’ as a reference, species-specific KASP primers were designed for PCR amplification. Based on the genotyping results, we constructed cultivar DNA fingerprints and assessed the efficiency of core SNP markers for cultivar identification. Molecular IDs for O. fragrans cultivars were established by integrating phenotypic information codes with molecular fingerprint codes.【Result】We retained a total of 14 core SNP loci from genomic SNPs that fully discriminated the sequenced cultivars. The PIC values of these loci ranged from 0.246 to 0.375, with an average of 0.335, and the He indices ranged from 0.288 to 0.500, averaging 0.431. The KASP primers designed for these core SNP loci produced accurate genotyping results, enabling us to construct DNA fingerprints capable of distinguishing all 90 tested cultivars, including those not previously sequenced. Each cultivar was assigned a molecular ID composed of 34 digits.【Conclusion】In conclusion, 14 core SNP loci (SNP1 to SNP14) were identified that effectively discriminate among at least 90 O. fragrans cultivars. Unique molecular ID codes were constructed using DNA fingerprint codes along with serial codes derived from cultivar group types and phenotypic characteristics. Finally, barcode and quick response (QR) codes were generated for each cultivar.

Key words

Osmanthus fragrans / cultivar identification / single nucleotide polymorphism (SNP) / phenotype / molecular ID

Cite this article

Download Citations
WANG Yihan , LIU Jiaojiao , JIN Peiquan , et al . Construction of molecular ID for Osmanthus fragrans cultivars based on phenotypic traits and single nucleotide polymorphisms (SNPs)[J]. JOURNAL OF NANJING FORESTRY UNIVERSITY. 2024, 48(4): 12-24 https://doi.org/10.12302/j.issn.1000-2006.202405026

References

[1]
赵宏波, 郝日明, 胡绍庆. 中国野生桂花的地理分布和种群特征[J]. 园艺学报, 2015, 42(9):1760-1770.
ZHAO H B, HAO R M, HU S Q. Geographic distribution and population characteristics of Osmanthus fragrans[J]. Acta Hortic Sin, 2015, 42(9):1760-1770.DOI: 10.16420/j.issn.0513-353x.2014-0939.
[2]
向其柏, 刘玉莲. 中国桂花品种图志[M]. 杭州: 浙江科学技术出版社, 2008.
XIANG Q B, LIU Y L. An illustrated monograph of the sweet osmanthus cultivars in China[M]. Hangzhou: Zhejiang Science & Technology Press, 2008.
[3]
杨康民. 中国桂花[M]. 北京: 中国林业出版社, 2013.
YANG K M. Chinese osmanthus[M]. Beijing: China Forestry Publishing House, 2013.
[4]
向民, 段一凡, 向其柏. 木犀属品种国际登录中心年报(1) 彩叶桂品种群的建立[J]. 南京林业大学学报(自然科学版), 2014, 38(1):2,187.
XIANG M, DUAN Y F, XIANG Q B. International Cultivar Registration Center for Osmanthus,Nanjing Forestry University.Establishment of a new group-Osmanthus fragrans Colour Group[J]. J Nanjing For Univ (Nat Sci Ed), 2014, 38(1):2,187.DOI: 10.3969/j.issn.1000-2006.2014.01.034.
[5]
高丽, 何岳球, 孙志国, 等. 三个 “新发展” 下咸宁市中国桂花城传承创新发展研究[J]. 安徽农业科学, 2023, 51(12):256-259.
GAO L, HE Y Q, SUN Z G, et al. Research on the inheritance,innovation and high-quality development of Chinese Osmanthus city in Xianning under three “new developments”[J]. J Anhui Agric Sci, 2023, 51(12):256-259.DOI: 10.3969/j.issn.0517-6611.2023.12.058.
[6]
刘燕培, 李书情, 王长海, 等. 桂花新品种‘潢川丹桂’[J]. 园艺学报, 2022, 49(S2):229-230.
LIU Y P, LI S Q, WANG C H, et al. A new Osmanthus fragrans cultivar ‘Huangchuan Dangui’[J]. Hortic Plant J, 2022, 49(S2):229-230. DOI: 10.16420/j.issn.0513-353x.2022-0376. (in Chinese with English abstract)
[7]
王良桂, 潘多, 丁卉芬, 等. 彩叶桂新品种‘南林彩云’[J]. 南京林业大学学报(自然科学版), 2023, 47(2):243-244.
WANG L G, PAN D, DING H F, et al. Osmanthus fragrans ‘Nanlin Caiyun’:a new cultivar of Osmanthus[J]. J Nanjing For Univ (Nat Sci Ed), 2023, 47(2):243-244.
[8]
国家质量监督检验检疫总局, 中国国家标准化管理委员会. 植物新品种特异性、一致性、稳定性测试指桂花:GB/T 24885—2010[S]. 北京: 中国标准出版社, 2011.
General Administration of Quality Supervision,Inspection and Quarantine of the People’s Republic of China, Standardization Administration of the People’s Republic of China. Guidelines for the conduct of test for distinctness,uniformity and stability-Sweet osmanthus(Osmanthus fragrans L.):GB/T 24885-2010[S]. Beijing: Standards Press of China, 2011.
[9]
UPOV. TGP/15. Guidance on the use of biochemical and molecular markers in the examination of distinctness,uniformity and stability(DUS). Geneva:UPOV, 2013.
[10]
李梅. 桂花种质资源遗传多样性研究及品种鉴定[D]. 南京: 南京农业大学, 2009.
LI M. Study on genetic diversity and cultivar identification of Osmanthus fragrans germplasm[D]. Nanjing: Nanjing Agricultural University, 2009.
[11]
邱帅, 吴光洪, 陈徐平, 等. 基于相关序列扩增多态性分子标记的桂花栽培品种演化分析[J]. 浙江大学学报(农业与生命科学版), 2017, 43(4):404-415.
QIU S, WU G H, CHEN X P, et al. Evolution analysis of sweet Osmanthus(Osmanthus fragrans) cultivars based on sequence-related amplified polymorphism molecular marker[J]. J Zhejiang Univ (Agric Life Sci), 2017, 43(4):404-415.DOI: 10.3785/j.issn.1008-9209.2016.08.241.
[12]
乔中全, 王晓明, 李永欣, 等. 桂花优良品种‘珍珠彩桂’遗传多样性的ISSR分析及指纹图谱构建[J]. 湖南林业科技, 2016, 43(3):1-5.
QIAO Z Q, WANG X M, LI Y X, et al. Genetic diversity and fingerprint construction of varieties of Osmanthus fragrans ‘Zhenzhu Caigui’ by ISSR markers[J]. Hunan For Sci Technol, 2016, 43(3):1-5.DOI: 10.3969/j.issn.1003-5710.2016.03.001.
[13]
罗仙英, 桂敬飞, 严治, 等. 贵州桂花种质资源遗传多样性的ISSR分析[J]. 种子, 2017, 36(6):62-66.
LUO X Y, GUI J F, YAN Z, et al. Genetic diversity analyses of Osmanthus fragrans lour.in Guizhou with ISSR markers[J]. Seed, 2017, 36(6):62-66.DOI: 10.16590/j.cnki.1001-4705.2017.06.062.
[14]
韩远记. 桂花品种资源遗传多样性的AFLP分析[D]. 开封: 河南大学, 2008.
HAN Y J. Study on the genetic diversity of Osmanthus fragrans cultivars by AFLP markers[D]. Kaifeng: Henan University, 2008.
[15]
YAN X Y, XIAO B L, HAN Y J, et al. AFLP analysis of genetic relationships and diversity of some Chinese Osmanthus fragrans cultivars[J]. Life Sci J, 2009, 6(2):11-16.
[16]
ZHANG Z R, FAN D M, GUO S Q, et al. Development of 29 microsatellite markers for Osmanthus fragrans (Oleaceae),a traditional fragrant flowering tree of China[J]. Am J Bot, 2011, 98(12):e356-e359.DOI: 10.3732/ajb.1100241.
[17]
DUAN Y F, WANG X R, XIANG Q B, et al. Genetic diversity of androdioecious Osmanthus fragrans (Oleaceae) cultivars using microsatellite markers[J]. Appl Plant Sci, 2013, 1(6): apps.1200092.DOI: 10.3732/apps.1200092.
[18]
李军, 董彬, 张超, 等. 桂花EST-SSR引物开发及在品种鉴定中的应用[J]. 浙江农林大学学报, 2018, 35(2):306-313.
LI J, DONG B, ZHANG C, et al. EST-SSR primers and their application in cultivar identification of Osmanthus fragrans[J]. J Zhejiang A F Univ, 2018, 35(2):306-313.DOI: 10.11833/j.issn.2095-0756.2018.02.015.
[19]
YANG X L, YUE Y Z, LI H Y, et al. The chromosome-level quality genome provides insights into the evolution of the biosynthesis genes for aroma compounds of Osmanthus fragrans[J]. Hortic Res, 2018, 5:72.DOI: 10.1038/s41438-018-0108-0.
[20]
CHEN H G, ZENG X L, YANG J, et al. Whole-genome resequencing of Osmanthus fragrans provides insights into flower color evolution[J]. Hortic Res, 2021, 8(1):98.DOI: 10.1038/s41438-021-00531-0.
[21]
LI Y, ZHAO H, XIA H X, et al. Multi-omics analyses provide insights into the genomic basis of differentiation among four sweet osmanthus groups[J]. Plant Physiol, 2024:kiae280.DOI: 10.1093/plphys/kiae280.
[22]
魏中艳, 李慧慧, 李骏, 等. 应用SNP精准鉴定大豆种质及构建可扫描身份证[J]. 作物学报, 2018, 44(3):315-323.
WEI Z Y, LI H H, LI J, et al. Accurate identification of varieties by nucleotide polymorphisms and establishment of scannable variety IDs for soybean germplasm[J]. Acta Agron Sin, 2018, 44(3):315-323.DOI: 10.3724/SP.J.1006.2018.000315.
[23]
DU H S, YANG J J, CHEN B, et al. Target sequencing reveals genetic diversity,population structure,core-SNP markers,and fruit shape-associated loci in pepper varieties[J]. BMC Plant Biol, 2019, 19(1):578.DOI: 10.1186/s12870-019-2122-2.
[24]
BUTTON P. The international union for the protection of new varieties of plants (upov) recommendations on variety denominations[J]. Acta Hortic, 2008(799):191-200.DOI: 10.17660/actahortic.2008.799.27.
[25]
HEO M S, HAN K, KWON J K, et al. Development of SNP markers using genotyping-by-sequencing for cultivar identification in rose (Rosa hybrida)[J]. Hortic Environ Biotechnol, 2017, 58(3):292-302.DOI: 10.1007/s13580-017-0268-0.
[26]
贾清香. 基于RAD测序的野鸢尾和射干的SSR及SNP特征分析[D]. 沈阳: 沈阳农业大学, 2018.
JIA Q X. Characteristics of SSR and SNP in I. dichotoma and I. domestica using RAD sequencing[D]. Shenyang: Shenyang Agricultural University, 2018.
[27]
MARTINA M, ACQUADRO A, PORTIS E, et al. Diversity analyses in two ornamental and large-genome Ranunculaceae species based on a low-cost Klenow NGS-based protocol[J]. Front Plant Sci, 2023, 14:1187205.DOI: 10.3389/fpls.2023.1187205.
[28]
SEMAGN K, BABU R, HEARNE S, et al. Single nucleotide polymorphism genotyping using Kompetitive Allele Specific PCR (KASP):Overview of the technology and its application in crop improvement[J]. Mol Breed, 2014, 33(1):1-14.DOI: 10.1007/s11032-013-9917-x.
[29]
HE C L, HOLME J, ANTHONY J. SNP genotyping: the KASP assay[M]. New York: Humana Press, 2014:75-86. DOI: 10.1007/978-1-4939-0446-4_7.
[30]
GANAL M W, WIESEKE R, LUERSSEN H, et al. High-throughput SNP profiling of genetic resources in crop plants using genotyping arrays[M]// Genom Plant Genet Resour. Dordrecht: Springer, 2014:113-130. DOI: 10.1007/978-94-007-7572-5_6.
[31]
DO KIM K, KANG Y N, KIM C. Application of genomic big data in plant breeding:past,present,and future[J]. Plants, 2020, 9(11):1454.DOI: 10.3390/plants9111454.
[32]
NGUYENen, TOAN K, HA S T T, et al. Analysis of Chrysanthemum genetic diversity by genotyping-by-sequencing[J]. Hortic Environ Biotechnol, 2020, 61:903-913. DOI: 10.1007/s13580-020-00274-2.
[33]
WANG F Q, FAN X C, ZHANG Y, et al. Establishment and application of an SNP molecular identification system for grape cultivars[J]. J Integr Agric, 2022, 21(4):1044-1057.DOI: 10.1016/S2095-3119(21)63654-7.
[34]
LI Z Y, YU H L, LI X, et al. Kompetitive allele-specific PCR (KASP) genotyping and heterotic group classification of 244 inbred lines in cabbage (Brassica oleracea L.var.capitata)[J]. Euphytica, 2020, 216(7):106.DOI: 10.1007/s10681-020-02640-8.
[35]
RASHEED A, WEN W E, GAO F M, et al. Development and validation of KASP assays for genes underpinning key economic traits in bread wheat[J]. Theor Appl Genet, 2016, 129(10):1843-1860.DOI: 10.1007/s00122-016-2743-x.
[36]
GREWAL S, HUBBART-EDWARDS S, YANG C Y, et al. Rapid identification of homozygosity and site of wild relative introgressions in wheat through chromosome-specific KASP genotyping assays[J]. Plant Biotechnol J, 2020, 18(3):743-755.DOI: 10.1111/pbi.13241.
[37]
李书情. 基于全基因组SNP的桂花品种精准鉴定体系的构建及应用[D]. 郑州: 河南农业大学, 2023.
LI S Q. Construction and application of accurate identification system of Osmanthus fragrans varieties based on whole-genome SNP[D]. Zhengzhou: Henan Agricultural University, 2023.DOI: 10.27117/d.cnki.ghenu.2023.000780.
[38]
POREBSKI S, BAILEY L G, BAUM B R. Modification of a CTAB DNA extraction protocol for plants containing high polysaccharide and polyphenol components[J]. Plant Mol Biol Report, 1997, 15(1):8-15.DOI: 10.1007/BF02772108.
[39]
CAI X, MAI R Z, ZOU J J, et al. Analysis of aroma-active compounds in three sweet Osmanthus (Osmanthus fragrans) cultivars by GC-olfactometry and GC-MS[J]. J Zhejiang Univ Sci B, 2014, 15(7):638-648.DOI: 10.1631/jzus.B1400058.
[40]
FU J X, HOU D, ZHANG C, et al. The emission of the floral scent of four Osmanthus fragrans cultivars in response to different temperatures[J]. Molecules, 2017, 22(3):430.DOI: 10.3390/molecules22030430.
[41]
HAN Y J, LI L X, DONG M F, et al. cDNA cloning of the phytoene synthase (PSY) and expression analysis of PSY and carotenoid cleavage dioxygenase genes in Osmanthus fragrans[J]. Biologia, 2013, 68(2):258-263.DOI: 10.2478/s11756-013-0002-z.
[42]
路飞. 桂花新品种DUS测试指南的研制[D]. 南京: 南京林业大学, 2009.
LU F. Study on the design of DUS testing guideline for new varieties of Osmanthus fragrans[D]. Nanjing: Nanjing Forestry University, 2009.
[43]
VOSS D H, HALE W N. A comparison of the three editions of the royal horticultural society colour chart[J]. Hortscience, 1998, 33:13-17.DOI: 10.21273/HORTSCI.33.1.13.
[44]
臧德奎, 向其柏. 中国桂花品种分类研究[J]. 中国园林, 2004, 20(11):40-49.
ZANG D K, XIANG Q B. Studies on the cultivar classification of Chinese sweet Osmanthus[J]. J Chin Landsc Archit, 2004, 20(11):40-49.DOI: 10.3969/j.issn.1000-6664.2004.11.011.
[45]
尚富德, 陈仲芳, 刘玉莲, 等. 桂花品种资源调查方法研究[J]. 河南大学学报(自然科学版), 2003, 33(1):9-13.
SHANG F D, CHEN Z F, LIU Y L, et al. Study on the investigation of cultivar resources of Osmanthus fragrans[J]. J Henan Univ (Nat Sci), 2003, 33(1):9-13.DOI: 10.15991/j.cnki.411100.2003.01.003.
[46]
GLEISER G, VERDÚ M. Repeated evolution of dioecy from androdioecy in Acer[J]. New Phytol, 2005, 165(2):633-640.DOI: 10.1111/j.1469-8137.2004.01242.x.
[47]
DANECEK P, AUTON A, ABECASIS G, et al. The variant call format and VCFtools[J]. Bioinformatics, 2011, 27(15):2156-2158.DOI: 10.1093/bioinformatics/btr330.
[48]
李梓榕, 袁雄, 陈叶, 等. 基于全基因组SNP高效鉴定水稻种质资源并构建指纹图谱[J]. 分子植物育种, 2020, 18(18):6050-6057.
LI Z R, YUAN X, CHEN Y, et al. Effective identification for varieties by genome-wide SNPs and establishment of fingerprint for rice germplasm[J]. Mol Plant Breed, 2020, 18(18):6050-6057.DOI: 10.13271/j.mpb.018.006050.
[49]
SHEN Y S, WANG J S, SHAW R K, et al. Development of GBTS and KASP panels for genetic diversity,population structure,and fingerprinting of a large collection of broccoli (Brassica oleracea L.var.italica) in China[J]. Front Plant Sci, 2021, 12:655254.DOI: 10.3389/fpls.2021.655254.
[50]
LIU K J, MUSE S V. PowerMarker:An integrated analysis environment for genetic marker analysis[J]. Bioinformatics, 2005, 21(9):2128-2129.DOI: 10.1093/bioinformatics/bti282.
[51]
杨康民, 朱文江. 桂花[M]. 上海: 上海科学技术出版社, 1999.
YANG K M, ZHU W J. Osmanthus fragrans[M]. Shanghai: Shanghai Scientific & Technical Publishers, 1999. (in Chinese)
[52]
任海龙, 许东林, 张晶, 等. 菜薹KASP-SNP指纹图谱构建及品种鉴定[J]. 园艺学报, 2023, 50(2):307-318.
REN H L, XU D L, ZHANG J, et al. Establishment of SNP fingerprinting and identification of Chinese flowering cabbage varieties based on KASP genotyping[J]. Acta Hortic Sin, 2023, 50(2):307-318.DOI: 10.16420/j.issn.0513-353x.2021-1046.
[53]
王富强, 樊秀彩, 张颖, 等. SNP分子标记在作物品种鉴定中的应用和展望[J]. 植物遗传资源学报, 2020, 21(5):1308-1320.
WANG F Q, FAN X C, ZHANG Y, et al. Application and prospect of SNP molecular markers in crop variety identification[J]. J Plant Genet Resour, 2020, 21(5):1308-1320.DOI: 10.13430/j.cnki.jpgr.20200309002.
[54]
LI H L, XIAO W J, TONG T, et al. The specific DNA barcodes based on chloroplast genes for species identification of Orchidaceae plants[J]. Sci Rep, 2021, 11(1):1424.DOI: 10.1038/s41598-021-81087-w.
[55]
樊晓静, 于文涛, 蔡春平, 等. 利用SNP标记构建茶树品种资源分子身份证[J]. 中国农业科学, 2021, 54(8):1751-1772.
FAN X J, YU W T, CAI C P, et al. Construction of molecular ID for tea cultivars by using of single-nucleotide polymorphism(SNP) markers[J]. Sci Agric Sin, 2021, 54(8):1751-1772.DOI: 10.3864/j.issn.0578-1752.2021.08.014.
[56]
赵仁欣, 李森业, 郭瑞星, 等. 利用SNP芯片构建我国冬油菜参试品种DNA指纹图谱[J]. 作物学报, 2018, 44(7):956-965.
ZHAO R X, LI S Y, GUO R X, et al. Construction of DNA fingerprinting for Brassica napus varieties based on SNP chip[J]. Acta Agron Sin, 2018, 44(7):956-965.DOI: 10.3724/SP.J.1006.2018.00956.
[57]
朱国忠, 张芳, 付洁, 等. 适于陆地棉品种身份鉴定的SNP核心位点筛选与评价[J]. 作物学报, 2018, 44(11):1631-1639.
ZHU G Z, ZHANG F, FU J, et al. Genome-wide screening and evaluation of SNP core loci for identification of upland cotton varieties[J]. Acta Agron Sin, 2018, 44(11):1631-1639.DOI: 10.3724/SP.J.1006.2018.01631.9.
PDF(5559 KB)

Accesses

Citation

Detail

Sections
Recommended
The full text is translated into English by AI, aiming to facilitate reading and comprehension. The core content is subject to the explanation in Chinese.

/