基于流式细胞术对朱砂根种质资源基因组大小的测定

康阳; 赵凯; 刘钰颖; 王钦; 涂松; 王睿昕; 朱云君; 刘梓富; 彭东辉

doi:10.19303/j.issn.1008-0384.2023.02.002

基于流式细胞术对朱砂根种质资源基因组大小的测定

doi: 10.19303/j.issn.1008-0384.2023.02.002

康阳^{1, 2,},
赵凯³,
刘钰颖^{1, 2},
王钦^{1, 2},
涂松^{1, 2},
王睿昕^{1, 2},
朱云君^{1, 2},
刘梓富⁴,
彭东辉^{1, 2, ,}

1.
福建农林大学风景园林学院，福建　福州　350002
2.
福建省观赏植物种质资源创新与应用工程技术研究中心，福建　福州　350002
3.
福建师范大学生命科学学院，福建　福州　350100
4.
武平县林业局，福建　武平　364300

基金项目: 福建省科技计划项目（2022L3008）；福建省林业科技推广项目(2021TG02)

详细信息

作者简介:
康阳（1997−），男，硕士研究生，研究方向：园林植物与应用（E-mail：1459666833@qq.com）

通讯作者:
彭东辉（1971-），男，博士，教授，研究方向：园林植物与应用（E-mail：fjpdh@126.com）

中图分类号: S686
计量
- 文章访问数: 468
- HTML全文浏览量: 245
- PDF下载量: 28
- 被引次数: 0
出版历程
- 收稿日期: 2022-08-25
- 修回日期: 2022-10-08
- 网络出版日期: 2023-03-05
- 刊出日期: 2023-02-28

Flow Cytometric Determination on Genome Size of Ardisia crenata Germplasms

KANG Yang^{1, 2
,},
ZHAO Kai³,
LIU Yuying^{1, 2},
WANG Qin^{1, 2},
TU Song^{1, 2},
WANG Ruixin^{1, 2},
ZHU Yunjun^{1, 2},
LIU Zifu⁴,
PENG Donghui^{1, 2
, ,}

1.
College of Landscape Architecture, Fujian Agriculture and Forestry University, Fuzhou, Fujian　350002, China
2.
Fujian Engineering Research Center of Ornamental Plant Germplasm Resources Innovation and Application, Fuzhou, Fujian　350002, China
3.
Fujian Normal University, Fuzhou, Fujian　350100, China
4.
Wuping County Forestry Bureau, Fujian　364300, China

摘要

摘要: 目的基于流式细胞术初步探索测定朱砂根基因组大小的方法和流程，为朱砂根基因组文库的建立、基因组全序列测定及其基因组学研究等工作的开展提供基础数据。方法以24份朱砂根（Ardisia crenata）种质资源为供试材料，包括22个人工选育品种和2个野生种质资源，并以番茄（Lycopersicon esculentum）作为内参样本，利用流式细胞术对朱砂根基因组大小进行测定。结果 24份朱砂根基因组大小（C值）为1.77～2.41 Gb，平均大小1.87 Gb；其中玛瑙红（Z-17）、霞珠（Z-20）、珠塔（Z-22） C值最小，均为1.77 Gb，赤丹 C最大，为2.41Gb，部分品种间基因组大小存在一定程度的差异。结论首次测定朱砂根种质资源的基因组大小，其研究结果可为朱砂根基因组文库的建立、基因组全序列测定及其基因组学研究等工作的开展提供基础数据。
- 朱砂根 /
- 流式细胞术 /
- 基因组大小 /
- 种质资源
Abstract: Objective Genome sizes of the economically, medicinally, and ornamentally valuable Ardisia crenata were determined using flow cytometry. Methods Genome sizes of 22 selectively bred cultivars and two wild A. crenata germplasms, along with Lycopersicon esculentum as internal reference, were determined by flow cytometry. Results The sizes ranged from 1.77 Gb to 2.41 Gb averaging 1.87 Gb with the smallest C-value of 1.77 Gb found on Agate Red (Z-17), Kasumi Pearl (Z-20), and Zhu Ta (Z-22), while the largest of 2.41 Gb on Chidan. Variations existed among the germplasms. Conclusion For the first time, the genome sizes of various A. crenata were determined to be available for library construction and genomics studies on the valuable natural resource.
- Ardisia crenata /
- flow cytometry /
- genome size /
- germplasms

HTML全文

图 1 流式细胞仪检测24份朱砂根荧光峰值

荧光峰值直方图左侧峰代表内参番茄，右侧峰代表朱砂根种质；FL2-H:荧光脉冲高度。

Figure 1. Fluorescence peak histogram of 24 A. crenata detected by flow cytometry

Left peak on fluorescence histogram represents internal reference L. esculentum; right peak, A. crenata varieties.FL2-H:Fluore scence pulse height.

下载: 全尺寸图片幻灯片

表 1 24份朱砂根供试材料采样信息

Table 1. Sampling of 24 A. crenata germplasms

编号 No.	种质名称 Germplasm name	种质类别 Germplasm type	编号 No.	种质名称 Germplasm name	种质类别 Germplasm type
Z-01	赤丹 Chi Dan	栽培品种 Cultivated variety	Z-13	福株 Fu Zhu	栽培品种 Cultivated variety
Z-02	锦绣 Jin Xiu	栽培品种 Cultivated variety	Z-14	金边富贵 Jin Bian Fu Gui	栽培品种 Cultivated variety
Z-03	绿翡翠 Green Jade	栽培品种 Cultivated variety	Z-15	金玉满堂 Jin Yu Man Tang	栽培品种 Cultivated variety
Z-04	粉佳人 Pink Beauty	栽培品种 Cultivated variety	Z-16	竹叶富贵 Bamboo Leaf Fu Gui	栽培品种 Cultivated variety
Z-05	平安富贵 Ping An Fu Gui	栽培品种 Cultivated variety	Z-17	玛瑙红 Onyx Red	栽培品种 Cultivated variety
Z-06	金冠 Golden Crown	栽培品种 Cultivated variety	Z-18	碧珠 Bi Zhu	栽培品种 Cultivated variety
Z-07	福满堂 Fu Man Tang	栽培品种 Cultivated variety	Z-19	龙珠 Long Zhu	栽培品种 Cultivated variety
Z-08	碧霞珠 Bixia Zhu	栽培品种 Cultivated variety	Z-20	霞珠 Xia Zhu	栽培品种 Cultivated variety
Z-09	金富贵 Jin Fu Gui	栽培品种 Cultivated variety	Z-21	仙桃 Xian Tao	栽培品种 Cultivated variety
Z-10	梁野富贵 Liang Ye Fu Gui	栽培品种 Cultivated variety	Z-22	珠塔 Zhu Ta	栽培品种 Cultivated variety
Z-11	赤玲珑 Red Lingerie	栽培品种 Cultivated variety	Z-(A)	朱砂根A A. crenata A	野生种质 Wild germplasm
Z-12	大富贵 Da Fu Gui	栽培品种 Cultivated variety	Z-(B)	朱砂根B A. crenata B	野生种质 Wild germplasm

下载: 导出CSV

表 2 流式细胞术测定的24份朱砂根品种资源基因组大小

Table 2. Genome sizes of 24 A. crenata determined by flow cytometry

样品编号Sample No.	内参荧光强度Internal reference fluorescence intensity	待测样品荧光强度Fluorescence intensity of the sample to be measured	比值Ratio	基因组大小Genome/Gb	样品编号Sample No.	内参荧光强度Internal reference fluorescence intensity	待测样品荧光强度Fluorescence intensity of the sample to be measured	比值Ratio	基因组大小Genome/Gb
Z-01	18.90	51.79	2.74	2.41	Z-13	26.61	63.08	2.37	2.09
Z-02	23.83	48.40	2.03	1.79	Z-14	27.09	65.13	2.40	2.12
Z-03	24.33	49.58	2.04	1.79	Z-15	27.22	64.87	2.38	2.10
Z-04	26.06	55.25	2.12	1.87	Z-16	26.68	55.70	2.09	1.84
Z-05	18.75	48.60	2.59	2.28	Z-17	26.63	53.53	2.01	1.77
Z-06	21.30	52.18	2.45	2.16	Z-18	27.19	55.46	2.04	1.79
Z-07	22.72	56.47	2.49	2.19	Z-19	27.61	55.73	2.02	1.78
Z-08	22.44	55.92	2.49	2.19	Z-20	28.26	56.70	2.01	1.77
Z-09	24.16	51.55	2.13	1.88	Z-21	28.56	58.22	2.04	1.79
Z-10	26.36	55.40	2.10	1.85	Z-22	28.34	56.89	2.01	1.77
Z-11	26.07	59.64	2.29	2.01	Z-(A)	28.77	58.62	2.04	1.79
Z-12	25.35	51.83	2.04	1.80	Z-(B)	28.54	58.29	2.04	1.80

下载: 导出CSV

参考文献(34)

[1]	张俊环, 杨丽, 姜凤超, 等. 基于流式细胞仪对杏属植物基因组大小的测定 [J]. 华北农学报, 2020, 35(5):32−38. ZHANG J H, YANG L, JIANG F C, et al. Estimation of genome size of apricots based on flow cytometry [J]. Acta Agriculturae Boreali-Sinica, 2020, 35(5): 32−38.(in Chinese)
[2]	ARUMUGANATHAN K, EARLE E D. Nuclear DNA content of some important plant species [J]. Plant Molecular Biology Reporter, 1991, 9(4): 415.
[3]	DOLEZEL J, BARTOS J, VOGLMAYR H, et al. Nuclear DNA content and genome size of trout and human[J]. Cytometry Part A: the Journal of the International Society for Analytical Cytology, 2003, 51(2): 127-128;authorreply129.
[4]	潘根, 董志雪, 唐蜻, 等. 流式细胞术测定玫瑰茄及红麻的基因组大小 [J]. 中国麻业科学, 2021, 43(5):217−221. PAN G, DONG Z X, TANG Q, et al. Estimation of genome size of two species from Hibiscus and kenaf by flow cytometry [J]. Plant Fiber Sciences in China, 2021, 43(5): 217−221.(in Chinese)
[5]	KANG M, TAO J J, WANG J, et al. Adaptive and nonadaptive genome size evolution in Karst endemic flora of China [J]. The New Phytologist, 2014, 202(4): 1371−1381. doi: 10.1111/nph.12726
[6]	李雯雯, 刘立强, 帕米尔·艾尼, 等. 利用流式细胞术鉴定新疆野杏染色体倍性和DNA含量 [J]. 农业生物技术学报, 2019, 27(3):542−550. LI W W, LIU L Q, PAMIER A N, et al. Identification of chromosomal ploidy and DNA content in Xinjiang Armeniaca vulgaris by flow cytometry [J]. Journal of Agricultural Biotechnology, 2019, 27(3): 542−550.(in Chinese)
[7]	孙绪, 姚成芬, 付思红, 等. 苗药朱砂根的HPLC指纹图谱研究 [J]. 中国药房, 2017, 28(30):4285−4288. SUN X, YAO C F, FU S H, et al. Study on HPLC fingerprint of Miao medicine Ardisia crenata [J]. China Pharmacy, 2017, 28(30): 4285−4288.(in Chinese)
[8]	叶晴, 陈金鹏, 凌悦, 等. 朱砂根化学成分和药理作用的研究进展 [J]. 中草药, 2022, 53(9):2851−2860. YE Q, CHEN J P, LING Y, et al. Research progress on chemical constituents and pharmacological effects of Ardisiae Crenatae Radix [J]. Chinese Traditional and Herbal Drugs, 2022, 53(9): 2851−2860.(in Chinese)
[9]	陈俊晖. 朱砂根品系评价和繁殖技术研究[D]. 福州: 福建农林大学, 2017. CHEN J H. Study on strains evaluate and propagation technique of Ardisia crenata[D]. Fuzhou: Fujian Agriculture and Forestry University, 2017. (in Chinese)
[10]	陈俊晖, 漆子钰, 骆亮, 等. 金边朱砂根组培快繁体系的建立 [J]. 江苏农业科学, 2017, 45(17):50−53. CHEN J H, QI Z Y, LUO L, et al. Establishment of tissue culture and rapid propagation system of Ardisia crenata in Phnom Penh [J]. Jiangsu Agricultural Sciences, 2017, 45(17): 50−53.(in Chinese)
[11]	周琪, 吕享, 林冰, 等. 朱砂根种胚繁育技术研究 [J]. 种子, 2022, 41(8):104−109,115. doi: 10.16590/j.cnki.1001-4705.2022.08.104 ZHOU Q, LYU X, LIN B, et al. Study on breeding regulation technology of Ardisia crenata Sims embryo [J]. Seed, 2022, 41(8): 104−109,115.(in Chinese) doi: 10.16590/j.cnki.1001-4705.2022.08.104
[12]	张建新, 郦枫, 马丽, 等. 镉胁迫下朱砂根和虎舌红生理响应及其镉抗性 [J]. 水土保持学报, 2017, 31(5):321−327. ZHANG J X, LI F, MA L, et al. Physiological responses and resistances of Ardisia crenata and a. mamillatato the treatments of cadmium stress [J]. Journal of Soil and Water Conservation, 2017, 31(5): 321−327.(in Chinese)
[13]	艾金祥, 宋嘉怡, 严浙楠, 等. 褪黑素对铅胁迫下虎舌红和朱砂根生理响应及DNA损伤的调控效应 [J]. 植物学报, 2022, 57(2):171−181. doi: 10.11983/CBB21191 AI J X, SONG J Y, YAN Z N, et al. Effects of exogenous melatonin on physiological response and DNA damage of Ardisia mamillata and A. crenata under lead stress [J]. Chinese Bulletin of Botany, 2022, 57(2): 171−181.(in Chinese) doi: 10.11983/CBB21191
[14]	骆亮, 张文春, 李龙, 等. 不同居群朱砂根(Ardisia crenata)的荧光ISSR遗传多样性分析 [J]. 分子植物育种, 2021, 19(18):6235−6247. LUO L, ZHANG W C, LI L, et al. Genetic diversity analysis of Ardisia crenata in different populations by fluorescence ISSR [J]. Molecular Plant Breeding, 2021, 19(18): 6235−6247.(in Chinese)
[15]	康阳, 刘梓富, 陈进燎, 等. 十六份朱砂根品种表型遗传多样性分析 [J]. 北方园艺, 2022(12):71−78. KANG Y, LIU Z F, CHEN J L, et al. Analysis of phenotypic genetic diversity of 16 Ardisia crenata cultivars [J]. Northern Horticulture, 2022(12): 71−78.(in Chinese)
[16]	刘雄伟, 刘畅, 曾宪法, 等. 朱砂根叶绿体全基因组解析及系统发育分析 [J]. 生物技术通报, 2023, 39(1):232−242. LIU X W, LIU C, ZENG X F, et al. Comparative and phylogenetic analyses of complete chloroplast genomes in Ardisia crenata [J]. Biotechnology Bulletin, 2023, 39(1): 232−242.(in Chinese)
[17]	刘畅, 潘婕, 刘雄伟, 等. 朱砂根AcGGPPS基因蛋白结构功能预测、密码子偏好性与进化分析[J/OL]. 分子植物育种, 2022: 1-24. (2022-03-03). https://kns.cnki.net/kcms/detail/46.1068.S.20220301.1351.012.html. LIU C, PAN J, LIU X W, et al. Structural and functional prediction, Codon preference and evolutionary analysis of AcGGPPS gene from Ardisia crenata Sims[J/OL]. Molecular Plant Breeding, 2022: 1-24. (2022-03-03). https://kns.cnki.net/kcms/detail/46.1068.S.20220301.1351.012.html.(in Chinese)
[18]	杨君. 朱砂根的转录组测序及与三萜皂苷合成相关基因的差异分析[D]. 雅安: 四川农业大学, 2015. YANG J. RNA-seq for DEG analysis about triterpenoid saponin synthesis and transcript profiling of Ardisia crenata Sims[D]. Yaan: Sichuan Agricultural University, 2015. (in Chinese)
[19]	田新民, 周香艳, 弓娜. 流式细胞术在植物学研究中的应用: 检测植物核DNA含量和倍性水平 [J]. 中国农学通报, 2011, 27(9):21−27. TIAN X M, ZHOU X Y, GONG N. Applications of flow cytometry in plant research—Analysis of nuclear DNA content and ploidy level in plant cells [J]. Chinese Agricultural Science Bulletin, 2011, 27(9): 21−27.(in Chinese)
[20]	DOLEŽEL J, BARTOŠ J. Plant DNA flow cytometry and estimation of nuclear genome size [J]. Annals of Botany, 2005, 95(1): 99−110. doi: 10.1093/aob/mci005
[21]	DOLEŽEL J, GREILHUBER J, SUDA J. Estimation of nuclear DNA content in plants using flow cytometry [J]. Nature Protocols, 2007, 2(9): 2233−2244. doi: 10.1038/nprot.2007.310
[22]	杜立颖, 冯仁青. 流式细胞术[M]. 2版. 北京: 北京大学出版社, 2014.
[23]	胡永乐, 宁书菊, 叶齐, 等. 流式细胞术测定马蓝基因组大小[J/OL]. 中成药, 2022: 1-3. (2022-05-25). https://kns.cnki.net/kcms/detail/31..1368.R20220524.1744.004.html. HU Y L, NING S J, YE Q, et al. Determination of Baphicacanthus cusia(Nees)Bremek genome size by flow cytometry [J/OL]. Chinese Traditional Patent Medicine, 2022: 1-3. (2022-05-25). https://kns.cnki.net/kcms/detail/31.1368.R.20220524.1744.004.html.(in Chinese)
[24]	金亮, 徐伟韦, 李小白, 等. DNA流式细胞术在植物遗传及育种中的应用 [J]. 中国细胞生物学学报, 2016, 38(2):225−234. JIN L, XU W W, LI X B, et al. Application of DNA flow cytometry to plant genetics and breeding [J]. Chinese Journal of Cell Biology, 2016, 38(2): 225−234.(in Chinese)
[25]	李春牛, 李先民, 黄展文, 等. 利用流式细胞术鉴定茉莉花基因组大小和染色体倍性 [J]. 热带作物学报, 2021, 42(5):1231−1236. LI C N, LI X M, HUANG Z W, et al. Genome size estimation and ploidy identification of Jasminum sambac by flow cytometry [J]. Chinese Journal of Tropical Crops, 2021, 42(5): 1231−1236.(in Chinese)
[26]	王利虎, 张琼, 陈凯, 等. 流式细胞术在植物倍性鉴定及基因组大小估测中的应用策略 [J]. 分子植物育种, 2021, 19(17):5833−5841. WANG L H, ZHANG Q, CHEN K, et al. Application strategy of flow cytometry in plant ploidy identification and genome size estimation [J]. Molecular Plant Breeding, 2021, 19(17): 5833−5841.(in Chinese)
[27]	THOMAS C A Jr. The genetic organization of chromosomes [J]. Annual Review of Genetics, 1971, 5: 237−256. doi: 10.1146/annurev.ge.05.120171.001321
[28]	李小东. 黄山常绿阔叶林树种基因组大小与表型分析[D]. 南京: 南京林业大学, 2019 LI X D. Genomic size and phenotype analysis of evergreen broad-leaved forest species in Huangshan Mountain[D]. Nanjing: Nanjing Forestry University, 2019. (in Chinese)
[29]	JORDAN G J, CARPENTER R J, KOUTOULIS A, et al. Environmental adaptation in stomatal size independent of the effects of genome size [J]. The New Phytologist, 2015, 205(2): 608−617. doi: 10.1111/nph.13076
[30]	CARTA A, PERUZZI L. Testing the large genome constraint hypothesis: Plant traits, habitat and climate seasonality in Liliaceae [J]. The New Phytologist, 2016, 210(2): 709−716. doi: 10.1111/nph.13769
[31]	BIÉMONT C. Genome size evolution: Within-species variation in genome size [J]. Heredity, 2008, 101(4): 297−298. doi: 10.1038/hdy.2008.80
[32]	张苏炯, 叶碧欢, 陈友吾, 等. 4种黄精属植物的基因组大小比较分析 [J]. 森林与环境学报, 2022, 42(2):193−198. ZHANG S J, YE B H, CHEN Y W, et al. Comparative analysis on genome sizes of four Polygonatum species [J]. Journal of Forest and Environment, 2022, 42(2): 193−198.(in Chinese)
[33]	石米娟, 程莹寅, 张婉婷, 等. 浅析基因组大小的进化机制 [J]. 科学通报, 2016, 61(30):3188−3195. doi: 10.1360/N972016-00728 SHI M J, CHENG Y Y, ZHANG W T, et al. The evolutionary mechanism of genome size [J]. Chinese Science Bulletin, 2016, 61(30): 3188−3195.(in Chinese) doi: 10.1360/N972016-00728
[34]	WOOD T E, TAKEBAYASHI N, BARKER M S, et al. The frequency of polyploid speciation in vascular plants [J]. Proceedings of the National Academy of Sciences of the United States of America, 2009, 106(33): 13875−13879. doi: 10.1073/pnas.0811575106