基于SRAP分子标记的流苏树天然群体遗传多样性研究

曲凯; 国浩平; 王宝锐; 周文玲; 侯丽丽; 李琴; 李际红; 程甜甜

doi:10.12171/j.1000-1522.20200212

基于SRAP分子标记的流苏树天然群体遗传多样性研究

1.
山东农业大学林学院，山东泰山森林生态系统国家定位研究站，黄河下游森林培育国家林业局重点实验室，山东泰安 271018
2.
山东方大工程有限责任公司嘉祥林场，山东济宁 272400
3.
泰安市泰山林业科学研究院，山东泰安 271000

基金项目: 山东省农业良种工程项目（2016LZGC036），山东省林业科技创新项目（LYCX02-2018-11），泰安市农业良种工程（2018）40号

详细信息

作者简介:
曲凯。主要研究方向：林木种质资源。Email：17661212306@163.com　地址：271000 山东省泰安市泰山区岱宗大街61号山东农业大学林学院

责任作者:
李际红，教授。主要研究方向：林木分子生物学及林木种质资源收集利用。Email：jhli@sdau.edu.cn　地址：同上

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出版历程
- 收稿日期: 2020-07-12
- 修回日期: 2020-08-14
- 网络出版日期: 2020-10-30
- 发布日期: 2021-01-06

Genetic diversity analysis of Chionanthus retusus natural population based on SRAP molecular markers

1.
College of Forestry, Shandong Agricultural University, Mountain Tai Forest Ecosystem Research Station of State Forestry and Grassland Administration, State Forestry and Grassland Administration Key Laboratory of Silviculture in Downstream Areas of The Yellow River, Taian 271018, Shandong, China
2.
Jiaxiang Forest Farm of Shandong Fangda Engineering Co., Ltd. Jining 272400, Shandong, China
3.
Taishan Forestry Science Institute, Taian 271000, Shandong, China

摘要

摘要:
目的揭示我国不同地区流苏树（Chionanthus retusus）天然群体的遗传多样性，更好地为合理保护和开发利用提供科学依据。
方法采用相关序列扩增多态性（SRAP）分子标记技术对不同地区的7个流苏树天然群体的62份样品进行了遗传多样性和群体遗传结构研究。
结果（1）7个流苏树天然群体具有较高的遗传多样性，8对SRAP引物共扩增出1 728条清晰条带，其中1 649条具有多态性，PPB（多态性条带比例）为95.43%；群体间的有效等位基因数为 1.213 7，Nei’s基因多样性指数为 0.153 7，Shannon’s信息多样性指数为0.268 0。（2）流苏树天然群体存在较高水平的种群内遗传变异和较低水平的群体间遗传变异（Gst = 0.133 6），7个流苏树天然群体间存在较高水平的基因交流（Nm = 3.243 7）。（3）流苏树群体间的遗传相似系数介于0.898 0 ~ 0.973 6之间，平均值为0.934 4，经Mantel检验（r = 0.288，P = 0.205）及群体间的聚类证明群体间的遗传距离与地理距离之间无明显相关性；62份流苏树初级种质聚类结果表明大部分种质表现为同一群体的多数个体聚在一起，部分种质存在不同群体间的个体聚在一起的现象，表现出群体间遗传变异相对稳定而种群内的遗传变异水平相对较高的特点，与基因多样性分析结果一致。
结论综合多因素分析推测，太行山地区可能是我国流苏树种质资源的主要产区。
- 流苏树 /
- SRAP /
- 遗传多样性 /
- 遗传结构
Abstract:
Objective This paper aims to reveal the genetic diversity of natural populations of Chionanthus retusus in different regions of China, and to provide a scientific basis for rational protection development and utilization.
Method The genetic diversity and population genetic structure of 62 samples from 7 Chionanthus retusus natural populations in different regions were studied using sequence related amplified polymorphism (SRAP) molecular marker technique.
Result Seven natural populations of Chionanthus retusus had higher genetic diversity, and 8 pairs of SRAP primers amplified a total of 1 728 clear bands, of which 1 649 were polymorphic, and the percentage of polymorphic bands (PPB) was 95.43%. The number of effective alleles between populations was 1.213 7, the diversity of Nei’s gene was 0.153 7, and the information diversity index of Shannon’s was 0.268 0. There were higher levels of intra-population genetic variation and lower levels of inter-population genetic variation among natural populations of Chionanthus retusus (Gst = 0.133 6), and higher levels of gene flow among seven natural populations of Chionanthus retusus (Nm = 3.243 7). The genetic similarity coefficient between Chionanthus retusus populations ranged from 0.898 0 to 0.973 6, with an average of 0.934 4. The Mantel test (r = 0.288, P = 0.205) and the clustering among populations proved that there was no significant correlation between genetic distance and geographical distance among populations. The clustering results of 62 primary germplasm showed that most of the germplasms were characterized by the fact that most individuals in the same population came together, and some germplasms had the phenomenon that individuals of different populations gathered together, showing that the genetic variation between populations was relatively stable and the level of genetic variation within the populations was relatively high, which was consistent with the results of genetic diversity analysis.
Conclusion Comprehensive multi-factor analysis speculated that Taihang Mountain area may be the main producing area of Chinese Chionanthus retusus germplasm resources.
- Chionanthus retusus /
- SRAP /
- genetic diversity /
- genetic structure

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图 1 不同流苏树天然群体间遗传关系聚类分析图

Figure 1. Dendrogram of UPGMA analysis of seven Chionanthus retusus natural populations

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图 2 7个流苏树天然群体间遗传距离与地理距离的相关性分析

Figure 2. Correlation analysis between genetic distance and geographic distance of seven Chionanthus retusus natural populations

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图 3 62份流苏树初级种质资源基于SRAP分析的UPGMA聚类结果

Figure 3. UPGMA dendrogram of primary collection of 62 Chionanthus retusus germplasm based on SRAP

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图 4 62份流苏树资源SRAP标记的主坐标分析图

Figure 4. Principal coordinate analysis for 62 samples of Chionanthus retusus

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表 1 流苏树天然群体采样地位置和生境

Table 1 Location and habitat of natural population sampling of Chionanthus retusus

群体及编号 Population and No.	取样株数 Sampling plant number	海拔 Altitude/m	纬度 Latitude (N)	经度 Longitude (E)
北京市怀柔区 Huairou District, Beijing City (B-H)	10	40	116°38′	40°17′
河北省保定市 Baoding City, Hebei Province (H-B)	5	20	115°28′	38°55′
河南省南阳市桐柏县 Tongbai County, Nanyang City, Henan Province (H-T)	10	240	113°17′	32°27′
山东省青州市 Qingzhou City, Shandong Province (S-Q)	10	250	118°18′	36°41′
山西省临汾市安泽县 Anze County, Linfen City, Shanxi Province (S-A)	10	260	112°14′	36°08′
江苏省宿迁市沭阳县 Shuyang County, Suqian City, Jiangsu Province(J-S)	10	10	118°39′	34°09′
湖北省安陆市 Anlu City, Hubei Province (H-A)	7	130	113°41′	31°15′

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表 2 7个流苏树天然群体间的地理距离

Table 2 Geographic distance of seven Chionanthus retusus natural populations km

群体 Population	H-T	H-A	S-A	S-Q	J-S	B-H	H-B
H-T	—
H-A	246	—
S-A	609	752	—
S-Q	856	966	672	—
J-S	698	795	799	373	—
B-H	1 039	1 204	736	512	809	—
H-B	862	993	547	448	710	221	—

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表 3 试验所用SRAP引物信息

Table 3 Information of SRAP primers used in the experiment

上游引物 Forward primer	引物序列 Primer sequence (5′→3′)	下游引物 Reverse primer	引物序列 Primer sequence (5′→3′)
T-ATA	TGAGTCCAAACCGGATA	G-AAT	GACTGCGTACGAATTAAT
T-AGC	TGAGTCCAAACCGGAGC	G-TGC	GACTGCGTACGAATTTGC
T-AAT	TGAGTCCAAACCGGAAT	G-GAC	GACTGCGTACGAATTGAC
T-ACC	TGAGTCCAAACCGGACC	G-TGA	GACTGCGTACGAATTTGA
T-AAG	TGAGTCCAAACCGGAAG	G-AAC	GACTGCGTACGAATTAAC
T-TAA	TGAGTCCAAACCGGTAA	G-GCA	GACTGCGTACGAATTGCA
T-ACA	TGAGTCCAAA CCGG ACA	G-CAA	GACTGCGTACG AATT CAA
T-TGT	TGAGTCCAAA CCGG TGT	G-AGC	GACTGCGTACG AATT AGC

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表 4 基于SRAP选择性扩增引物产生的条带多态性

Table 4 Polymorphism of SRAP bands obtained by selective amplification based on the primer combinations

引物组合 Primer combination	总带数 Total number of band	多态性条带数 Polymorphic band number	多态性条带比例 Percentage of polymorphic band/%
T-AGC/G-AAT	216	202	93.52
T-AGC/G-GCA	216	203	93.98
T-AGC/G-CAA	216	206	95.37
T-ACC/G-AAT	216	212	98.15
T-TAA/G-AAT	216	206	95.37
T-TAA/G-GCA	216	211	97.69
T-TAA/G-CAA	216	214	99.07
T-ACA/G-TGA	216	195	90.28
合计 Sum	1 728	1 649	—
平均 Mean	216	206.13	95.43

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表 5 基于不同引物组合的流苏树遗传多样性水平

Table 5 Genetic diversity level of Chionanthus retusus based on different primer combinations

引物组合 Primer combination	有效等位基因数 Number of effective allele (Ne)	Nei’s基因多样性指数 Nei’s gene diversity (H)	Shannon多态性信息指数 Shannon polymorphism information index (I)
T-AGC/G-AAT	1.216 6	0.151 6	0.261 1
T-AGC/G-GCA	1.235 6	0.163 3	0.278 4
T-AGC/G-CAA	1.237 4	0.163 9	0.277 8
T-ACC/G-AAT	1.227 3	0.164 8	0.286 7
T-TAA/G-AAT	1.162 6	0.122 7	0.224 4
T-TAA/G-GCA	1.221 2	0.168 5	0.298 7
T-TAA/G-CAA	1.215 9	0.159 0	0.280 5
T-ACA/G-TGA	1.193 2	0.135 6	0.236 3
平均 Mean	1.213 7	0.153 7	0.268 0

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表 6 7个流苏树天然群体内遗传多样性水平和显著性分析

Table 6 Analysis of genetic diversity and significance of the seven Chionanthus retusus natural populations

群体 Population	Ne	H	I
B-H	1.220 4a	0.146 1ab	0.238 9a
H-B	1.204 3a	0.131 5b	0.207 7b
H-T	1.195 0a	0.134 2ab	0.224 1ab
S-Q	1.206 1a	0.138 5ab	0.228 2ab
S-A	1.210 3a	0.143 7ab	0.238 9a
J-S	1.226 5a	0.150 4a	0.245 2a
H-A	1.207 4a	0.137 9ab	0.223 7ab
注：同列不同小写字母表示种群间差异显著(P < 0.05)。 Note: different lowercase letters in same column indicate significant differences among populations (P < 0.05).

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表 7 7个流苏树天然群体遗传分化分析

Table 7 Genetic differentiation of the seven Chionanthus retusus natural populations

所有群体 All population	总基因多样性指数 Total gene diversity index (Ht)	群体内基因多样性 Genetic diversity within the population (Hs)	群体间基因多样性 Genetic diversity between populations (Dst)	基因分化系数 Coefficient of gene differentiation (Gst)	基因流 Gene flow (Nm)
平均数 Mean	0.408 6	0.354 0	0.054 6	0.133 6	3.243 7
标准差 Standard deviation	0.027 6	0.012 0	—	—	—

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表 8 基于SRAP检测的7个流苏树天然群体间遗传一致度和遗传距离

Table 8 Genetic identity and genetic distance between seven Chionanthus retusus natural populations based on SRAP

群体 Population	H-T	H-A	S-A	S-Q	J-S	B-H	H-B
H-T	—	0.958 3	0.973 6	0.965 8	0.920 0	0.918 9	0.907 4
H-A	0.042 6	—	0.964 5	0.951 7	0.911 0	0.909 9	0.895 1
S-A	0.026 7	0.036 1	—	0.960 6	0.931 0	0.933 2	0.898 0
S-Q	0.034 8	0.049 5	0.040 2	—	0.954 7	0.947 2	0.935 8
J-S	0.083 4	0.093 3	0.071 5	0.046 4	—	0.957 8	0.904 1
B-H	0.084 5	0.094 4	0.069 2	0.054 2	0.043 1	—	0.922 8
H-B	0.097 2	0.110 8	0.107 5	0.066 3	0.100 9	0.080 4	—
注：右上部为遗传一致度，左下部为遗传距离。Notes: Nei’s genetic identity is showed above diagonal and genetic distance is showed below diagonal.

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