南京椴群体遗传多样性和遗传结构分析

doi:10.12302/j.issn.1000-2006.202110049

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南京林业大学学报（自然科学版） ›› 2023, Vol. 47 ›› Issue (1): 145-153.doi: 10.12302/j.issn.1000-2006.202110049

南京椴群体遗传多样性和遗传结构分析

王欢利(), 严灵君, 黄犀, 王仲伟, 汤诗杰()

江苏省中国科学院植物研究所,江苏省植物资源研究与利用重点实验室,江苏南京 210014

收稿日期:2021-10-25 接受日期:2021-11-29 出版日期:2023-01-30 发布日期:2023-02-01
通讯作者: 汤诗杰
基金资助:
国家自然科学基金青年基金项目(31700477);江苏省科技项目基础研究计划(自然科学基金)青年基金项目(BK20170619);江苏省林业科技创新与推广项目(LYKJ[2019]06)

Genetic diversity and genetic structure of Tilia miqueliana population

WANG Huanli(), YAN Lingjun, HUANG Xi, WANG Zhongwei, TANG Shijie()

Institute of Botany Jiangsu Province and Chinese Academy of Science,Jiangsu Key Laboratory for the Research and Utilization of Plant Resources, Nanjing 210014, China

Received:2021-10-25 Accepted:2021-11-29 Online:2023-01-30 Published:2023-02-01
Contact: TANG Shijie

摘要/Abstract

摘要： 【目的】 南京椴(Tilia miqueliana)为江苏省重要的乡土树种,野外资源稀缺。南京椴野外群体遗传多样性和遗传结构的探索,可为资源保护、品种选育及遗传改良提供依据。【方法】 以南京椴5个天然群体[江苏宝华山(P1)、牛首山(P2)、安徽皇藏峪(P3)、安徽蜀山(P4)和浙江天台山(P5)]93个体为实验材料,选用15对多态性EST-SSR引物,进行遗传多样性及群体遗传结构分析。【结果】 ①用15对引物共检测等位基因数(A)总和为96,平均值为6.4,四倍体基因型(G)和四倍体特异基因型(G_i)总和分别为441和251,特异基因型比率(R₁)和种质鉴别率(R₂)均值分别为45.73%和17.99%。②在5个群体中,每个位点等位基因数(A_loc)和四倍体基因型丰富度均值(G_loc)分别为5.50±2.43和9.41±4.29;平均观测杂合度(H_o)和平均期望杂合度(H_e)为0.61±1.43和0.62±0.14。参考各群体G_loc和H_e值,确定遗传多样性较高的群体为P1和P3。③群体间遗传分化较小,遗传分化系数(G_st)仅为0.030;AMOVA分子变异分析显示,群体多样性水平变异来自于群体内(96%)。④聚类和遗传结构Structure分析显示,5个群体可划分为2组(组1包括P1、P2和P5;组2包括P3和P4)。Mental检验结果表明遗传距离与地理距离之间无显著相关。【结论】 南京椴群体均具有丰富的遗传多样性,其中宝华山群体和皇藏峪群体多样性较高,群体扩张可能是以这两个种群为中心,经人类活动迁移至其他区域。但南京椴群体间未形成明显分化,主要是由于植株寿命长,群体缺乏自然更新,加之群体间存在人为种子传播。因此,本研究提出通过建立隔离区,明确优先保护群体、加大植株异交,并采用人工繁育及种质回迁的方式保护南京椴野外群体。

关键词: 南京椴, SSR标记, 遗传结构, 遗传多样性

Abstract:

【Objective】 Tilia miqueliana is an important native tree species in Jiangsu Province, and its wild germplasm resources are particularly scarce. Exploration of the genetic diversity and the genetic structure of T. miqueliana in the field population provides the scientific basis for its resource protection, variety breeding and genetic improvement.【Method】 Fifteen polymorphic EST-SSR primers were used to analyze the genetic diversity and population structure of 93 individuals from five natural populations of T. miqueliana located on Baohua Mountain (Jiangsu Province, P1), Niushou Mountain (Jiangsu Province, P2), Huangcangyu Nature Reserve (Anhui Province, P3), Shu Mountain (Anhui Province, P4) and Tiantai Mountain (Zhejiang Province, P5). 【Result】 (1) A total of 96 alleles (A) were detected using 15 EST-SSR primer pairs. The average allelic richness at a locus was 6.4. The total number of tetraploid genotypic richness (G) and tetraploid specific genotypes (G_i) were 441 and 251, respectively. The average of the specific genotype ratio (R₁) and the germplasm identification rate (R₂) were 45.73% and 17.99%, respectively. (2) In the five populations, the mean number of alleles per locus (A_loc) and the tetraploid genotypic richness (G_loc) were 5.50 ± 2.43 and 9.41 ± 4.29, respectively. The mean observed heterozygosity (H_o) and the mean expected heterozygosity (H_e) were 0.61 ± 1.43 and 0.62 ± 0.14, respectively. Based on the values for G_loc and H_e, the P1 and P3 populations were identified as having high genetic diversity. (3) The genetic differentiation coefficient (G_st) was 0.030, indicating a low genetic differentiation among the five populations. Analysis of molecular variance (AMOVA) showed that the variation in genetic diversity predominantly came from within the population (96%). (4) Cluster analysis and genetic structure analysis by structure showed that the five tested populations could be divided into two groups (Group1. P1, P2 and P5; Group2. P3 and P4). The Mantel test results showed that there was no significant correlation between the genetic differentiation and the geographic distance. 【Conclusion】 High levels of genetic diversity were found within the five populations of T. miqueliana, among which Baohua Mountain population and Huangcangyu Nature Reserve population showed considerably higher genetic diversity than that the other populations showed. Combined with the results from the clustering and genetic structure analysis, we can infer that population expansion may have been centered on these two populations and migrated to other regions through human activities. There was no pronounced genetic differentiation among the different geographic populations, which may be due to the long life of individuals, the lack of natural regeneration of populations, and seed dispersal among populations through human activities. Therefore, we propose to establish isolation areas, identify priority populations for protection, increase plant outcrossing, and use artificial breeding and germplasm regeneration to protect the wild populations.

Key words: Tilia miqueliana, SSR marker, genetic structure, genetic diversity

中图分类号:

S722

王欢利,严灵君,黄犀,等. 南京椴群体遗传多样性和遗传结构分析[J]. 南京林业大学学报（自然科学版）, 2023, 47(1): 145-153.

WANG Huanli, YAN Lingjun, HUANG Xi, WANG Zhongwei, TANG Shijie. Genetic diversity and genetic structure of Tilia miqueliana population[J].Journal of Nanjing Forestry University (Natural Science Edition), 2023, 47(1): 145-153.DOI: 10.12302/j.issn.1000-2006.202110049.

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种群编号 population No.	采集地点 location of collection	东经/ (°) longitude (E)	北纬/ (°) latitude (N)	个体数 sample size
P1	江苏宝华山 Baohua Mountain, Jiangsu	119.08	32.13	15
P2	江苏牛首山 Niushou Mountain, Jiangsu	118.75	31.90	16
P3	安徽皇藏峪 Huangcangyu, Anhui	117.07	34.02	33
P4	安徽蜀山 Shu Mountain, Anhui	117.18	31.84	17
P5	浙江天台山 Tiantai Mountain, Zhejiang	121.03	29.16	12

编号 No.	引物序列(5'→3') primer squence	产物长度/bp length of products	退火温度/℃ annealing tempreature	荧光标记 flourescent label
B1045	F: ACGGCTCCCTTATCACACTG R: TGCCATCTCACGTAGCTTTG	280	56.5	5'-FAM
B2405	F: TCTGCAAAGGAACTCTTCATCA R: TGGATTGCTAGTTGTCAAGCAT	218	54.0	5'-HEX
B2410	F: TATAGATGCCCGTTCCAAGC R: CATCAAGACAAGGGCAATGA	195	52.5	5'-HEX
B2550	F: ATTGTGCTGTGGTGGCATTA R: TACCCCTGCATGTTTTCCAT	226	53.0	5'-HEX
B2915	F: CGACCTGGTTCTCTGGTTTC R: CGACCAGGTGCATATAGCCT	268	54.5	5'-FAM
B485	F: TGGATGGTGATGGTAGGGTT R: GTGTGTCAACTGGCAGCAAT	278	54.0	5'-FAM
B75	F: GGAGAGAGAAAGGCAGAGCA R: TAAGGTGGTGATGACGTTGG	189	54.5	5'-TAMRA
B90	F: AGAGCAGAGCCGTTGGATAA R: GGAGAGAGAAAGGCAGAGCA	193	54.0	5'-FAM
C1050	F: ACCTGGAGGCTCTGGACTT R: ACACAGAGGTTCGGAAAATCG	259	54.0	5'-FAM
C2155	F: CGGTGTGTCGACATTGTTTC R: CTCTGGAAGAAGGGAACACG	219	53.5	5'-HEX
C2700	F: GCCAGCCATGTTTTGTTTCT R: CGTTCAATGGCCTCTTCATT	181	58.0	5'-FAM
C3235	F: GTTAGGAGCAATGATGCCGT R: CCACAGTCGCATCTTCTTCA	162	57.0	5'-FAM
E5	F: TTGCTGTTTCATTCTGCTGG R: AGTTTCGAATAGCGCCTTGA	178	53.0	5'-TAMRA
Tc11	F:AGCTATGAAAGAACTATCAAGAGAAAG R: CCCCAAGACATTGCAGTAGAAC	146	55.0	5'-TAMRA
Tc8	F: CGAAGAAACTGTCAAAACAACG R: AGCTGGGTTTTAGAGGATAGGG	160	58.0	5'-FAM

引物名称 primer	A	A_i	G	G_i	R₁/%	R₂/%	C_PIC	I
C1050	4	2	13	7	53.85	7.53	0.660 2	1.456 7
C2155	4	3	13	2	15.38	2.15	0.814 0	2.015 6
C2700	3	2	13	3	23.08	3.23	0.801 2	1.968 8
C3235	12	3	62	42	67.74	45.16	0.978 8	3.997 7
E05	4	2	15	4	26.67	4.30	0.829 5	2.189 8
Tc8	3	2	8	2	25.00	2.15	0.740 9	1.557 4
Tc11	6	2	33	13	39.39	13.98	0.943 7	3.162 9
B2250	6	2	21	8	38.10	8.60	0.888 4	2.569 1
B1045	4	2	18	5	27.78	5.38	0.888 4	2.472 9
B485	10	3	65	50	76.92	53.76	0.969 4	3.941 7
B2405	7	3	49	30	61.22	32.26	0.968 4	3.667 9
B75	9	3	43	28	65.12	30.11	0.914 1	3.185 3
B291 5	6	2	29	13	44.83	13.98	0.894 7	2.789 1
B2410	9	2	31	19	61.29	20.43	0.820 9	2.562 6
B90	9	3	42	25	59.52	26.88	0.952 7	3.387 8
总计total	96	—	441	251	—	—	—	—
均值mean	6.4	2.3	30.3	17	45.73	17.99	0.871 0	2.728 4

群体 population	N	A_l	A_loc	G_loc	H_o	H_e
P1	15	5.73±2.52	2.35±0.43	9.60±3.85	0.64±0.15	0.65±0.11
P2	16	5.27±2.43	2.34±0.38	8.33±4.66	0.63±0.16	0.63±0.15
P3	33	5.87±2.67	2.25±0.43	13.20±6.54	0.60±0.14	0.62±0.14
P4	17	5.20±2.54	2.23±0.44	9.20±3.14	0.59±0.15	0.61±0.14
P5	12	5.00±2.00	2.28±0.51	6.73±3.28	0.59±0.20	0.58±0.19
均值mean	18.6	5.50±2.43	2.29±0.44	9.41±4.29	0.61±1.43	0.62±0.14

引物名称 primer name	H_t	H_s	D_st	G_st
C1050	0.433	0.385	0.048	0.110
C2155	0.628	0.621	0.007	0.012
C2700	0.579	0.578	0.001	0.002
C3235	0.778	0.769	0.009	0.011
E5	0.597	0.567	0.030	0.051
Tc8	0.521	0.515	0.006	0.011
Tc11	0.686	0.669	0.017	0.024
B2250	0.514	0.491	0.023	0.045
B1045	0.681	0.647	0.034	0.050
B485	0.841	0.811	0.030	0.036
B2405	0.788	0.772	0.016	0.020
B75	0.756	0.746	0.010	0.013
B2915	0.540	0.520	0.020	0.036
B2410	0.425	0.434	0.011	0.026
B90	0.740	0.739	0.001	0.002
均值mean	0.634	0.616	0.018	0.030

南京椴群体遗传多样性和遗传结构分析

Genetic diversity and genetic structure of Tilia miqueliana population

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变异来源 source of variation	自由度 df	平方和 sum of square	变异组成 variance component	变异百分比/% percentage of variation	P_hi_PT
群体间 among populations	4	91.206	0.557	4	0.042
群体内 within populations	88	1 130.515	12.847	96
总变异 total	92	1 221.720	13.404	100

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