刺槐群体遗传多样性分析及其核心种质初步筛选

doi:10.11707/j.1001-7488.LYKX20240653

摘要/Abstract

摘要：

目的: 基于荧光SSR分子标记技术分析刺槐群体遗传多样性和遗传结构，初步筛选核心种质资源，为科学管理和保存刺槐种质资源、精确构建核心种质资源库以及有效提高育种效率提供理论参考。方法: 利用17对荧光SSR引物，分析来自美国阿巴拉契亚、北京、河南、山东、山西、辽宁6个种源地323份刺槐种质资源群体的遗传多样性和遗传结构，应用Gene Marker 2.2.0 结合POPGENE 32软件计算观测等位基因数（N_a）、有效等位基因数（N_e）、观测杂合度（H_o）、期望杂合度（H_e）、Shannon信息指数（I）、Nei’s基因多样性指数（H）和种群间F值。运用PowerMaker软件获得多态性信息含量（PIC）值。利用Structure软件分析刺槐种群结构，使用Python 3算法中的Evanno2005方法推断划分最佳遗传类群。采用最小距离逐步抽样法（LDSS）构建核心种质库后进行优化，t检验验证优化核心种质库的代表性。结果: 323份刺槐种质资源群体共扩增出135个等位基因，平均每对引物扩增7.941个等位基因，有效等位基因数均值2.808，Shannon信息指数均值1.200，观测杂合度均值0.191，期望杂合度均值0.588，Nei’s基因多样性指数均值0.589，多态信息含量均值0.544，表明刺槐群体遗传多样性较高；6个地理来源刺槐群体整体分析显示遗传多样性最高为辽宁、最低为北京；6个地理来源323份刺槐种质资源可划分为2个亚群，基因变异主要存在于种质资源个体间。最小距离逐步抽样法确定初始核心种质资源库最佳比例为20%，优化核心种质资源库比例为23.220%，确定248份保留种质、63份初始核心种质、12份补充种质，共筛选出75份核心种质，北京6份、河南8份、美国阿巴拉契亚17份、山西8份、山东29份、辽宁7份；核心种质资源库相比原始种质资源库N_a保留比例100.00%，N_e、I、H_o、H_e和H分别增加0.423、0.245、0.267、0.192和0.197，t检验表明核心种质库能够充分代表原始种质资源群体的遗传多样性。结论: 323份刺槐种质资源群体遗传多样性处于较高水平，遗传结构与地理来源相关性较小，初步构建的核心种质库可为下一步的资源保护和高效利用提供参考。

关键词: 刺槐, 荧光SSR, 遗传多样性, 遗传结构, 核心种质库

Abstract:

Objective: The genetic diversity and genetic structure of Robinia pseudoacacia populations were studied by fluorescent SSR molecular marker technique, and the core germplasm resources were preliminarily screened, in order to provide some theoretical references for scientific management and conservation of R. pseudoacacia germplasm resources, accurate construction of the core germplasm resource bank and effective improvement of breeding efficiency. Method: The genetic diversity and genetic structure of 323 R. pseudoacacia germplasm populations from 6 provenance regions of Appalachia in USA, Beijing, Henan, Shandong, Shanxi and Liaoning were analyzed using 17 pairs of fluorescent SSR primers. The software Gene Marker 2.2.0 and POPGENE32 were used to calculate and observed allele number (N_a), effective allele number (N_e), observed heterozygosity (H_o), expected heterozygosity (H_e), Shannon information index (I), Nei’s gene diversity index (H) and inter-population F value. The polymorphism information content (PIC) value was obtained by PowerMaker software. The software Structure was used to analyze the population structure of R. pseudoacacia, and the Evanno2005 method in Python 3 algorithm was used to infer and divide the best genetic groups. The least distance step-up sampling method (LDSS) was used to construct the core germplasm and optimize it. The representativeness of the optimized core germplasm bank was verified by t test. Result: A total of 135 alleles were amplified from 323 R. pseudoacacia germplasm resources. Through calculation and analysis, the results showed that the average value of allele number (N_a) amplified by each primer pair was 7.941, the average value of effective allele number (N_e) was 2.808, the average value of Shannon information index (I) was 1.200, the average value of observed heterozygosity (H_o) was 0.191, the average value of expected heterozygosity (H_e) was 0.588, the average value of Nei’s diversity index (H) was 0.589, and the average value of polymorphism information content (PIC) was 0.544, indicating that the genetic diversity of R. pseudoacacia population was high. According to the analysis of 6 geographical source populations of R. pseudoacacia, the highest genetic diversity was in Liaoning and the lowest was in Beijing. There were 6 geographical sources of 323 R. pseudoacacia germplasm resources, which could be divided into 2 subgroups, and the gene variation mainly existed among germplasm resources individuals. The LDSS was used to determine the optimal proportion of the initial core germplasm collection of 20% and the optimized core germplasm collection of 23.220%. After optimizing the core germplasm resource collection, 248 retained germplasm, 63 initial core germplasm and 12 supplementary germplasm were identified. A total of 75 core germplasm were screened, including 6 in Beijing, 8 in Henan, 17 in Appalachia of USA, 8 in Shanxi, 29 in Shandong and 7 in Liaoning. The retention rate of N_a was 100.00%, and N_e, I, H_o, H_e and H increased by 0.423, 0.245, 0.267, 0.192 and 0.197, respectively. The t test showed that the core germplasm collection could fully represent the genetic diversity of the original germplasm resource populations. Conclusion: The genetic diversity of 323 R. pseudoacacia germplasm resources is at a high level, and there is no significant correlation between genetic structure and geographical origin. The preliminarily constructed core germplasm collection can provide a reference for further resource conservation and efficient utilization.

Key words: Robinia pseudoacacia, fluorescent SSR, genetic diversity, genetic structure, core germplasm collection

中图分类号:

S792.26

梁燕,王因花,燕丽萍,孔雨光,李庆华,仲伟国,田泽新,董元夫,张元帅. 刺槐群体遗传多样性分析及其核心种质初步筛选[J]. 林业科学, 2025, 61(11): 242-254.

Yan Liang,Yinhua Wang,Liping Yan,Yuguang Kong,Qinghua Li,Weiguo Zhong,Zexin Tian,Yuanfu Dong,Yuanshuai Zhang. Analysis of Genetic Diversity in Robinia pseudoacacia Populations and Preliminary Screening of the Core Germplasm[J]. Scientia Silvae Sinicae, 2025, 61(11): 242-254.

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引物 Primer	重复单元 Repeat motifs	引物序列 Primer sequence		荧光类型 Fluorescence type
MQ7	(AAT)₁₁	F:	AACACGAGTCGAGGCTATCAAT	TARMRA
MQ7	(AAT)₁₁	R:	ACGGCTAGAGTCATTGGTTCAA	TARMRA
MQ12	(AT)₈	F:	AGAAAAGCGTCCATTGTCTAAT	FAM
MQ12	(AT)₈	R:	AGAGGGACAAGACAGAAAATG	FAM
MQ32	(TG)₁₀	F:	TCCCTCTCTTTCCGTTGATGAT	FAM
MQ32	(TG)₁₀	R:	AATCCTGGACGTGAAGAATAAA	FAM
MQ37	(TGT)₁₀	F:	ACGAATTCTTTCTTCCGCAAAA	HEX
MQ37	(TGT)₁₀	R:	TCGCCCTCTATTTCTTGTTCTT	HEX
MQ39	(A)₁₂	F:	CTTCGTGGTGGTGTCCTTTATA	TARMRA
MQ39	(A)₁₂	R:	CGAAAGAATCCACTGCCAATTT	TARMRA
MQ47	(TCA)₁₀	F:	TGGTTCCGTTGCTTTCTTATTC	ROX
MQ47	(TCA)₁₀	R:	AGACTATGCCTACGCCATATAA	ROX
MQ49	(T)₁₀	F:	ACTGCCGCCACTTATCTTTATT	FAM
MQ49	(T)₁₀	R:	GAACCGCTGGATCATCACTTAT	FAM
MQ54	(ACC)₁₀	F:	CAGTCCAAGGGTTGAAGTAAAA	HEX
MQ54	(ACC)₁₀	R:	GTCGTTAAACCCGGCAAAATA	HEX
MQ59	(T)₁₀	F:	AAATCGTCGAGAACCCTTTAAA	ROX
MQ59	(T)₁₀	R:	TTCTGGGCCGAGAGAATTATTT	ROX
MQ64	(T)₁₄	F:	GGATAGAGATATCGCCCGTTAT	TARMRA
MQ64	(T)₁₄	R:	TCGTGTGGATCAAAAGAAAGAT	TARMRA
CC2-48	(AGA)₁₁	F:	GACACTGAAATAGGCTCCTGAT	TARMRA
CC2-48	(AGA)₁₁	R:	ATTTTGGCAGGAGGGGAAATTA	TARMRA
CC2-54	(AAT)₁₀	F:	TCAGTCCAAGGGTTGAAGTAAA	HEX
CC2-54	(AAT)₁₀	R:	GTCGTTAAACCCGGCAAAATA	HEX
CC2-61	(A)₁₀	F:	TGAGGCCCCATATTGACTATCA	FAM
CC2-61	(A)₁₀	R:	TCGGGAGTCACAAATTCGTTAA	FAM
MQ16	(T)12c(A)₁₀	F:	ACCCTTACGCTTTGCAGATATA	FAM
MQ16	(T)12c(A)₁₀	R:	TCAACGTCCAATTTTCGGTAGA	FAM
RPly02	(CCA)₆	F:	TGTGAATGGTTGGTGGACAT	FAM
RPly02	(CCA)₆	R:	CGTTGCTTGGAGGAGAATAA	FAM
RPly22	(ACCTGA)₃	F:	ATCACATCTGTTCCTCCAC	TARMRA
RPly22	(ACCTGA)₃	R:	TTCTCCTCAGCCACTTCTTT	TARMRA
RPly109	(AG)₁₇	F:	GAGGAATCACAAAACCGTTTGG	HEX
RPly109	(AG)₁₇	R:	TGGGATTTGAGAGAGTGGTGGTG	HEX

位点Locus	N_a	N_e	I	H_o	H_e	H	PIC	F_st
MQ49	6.000	4.261	1.588	0.016	0.765	0.767	0.730	0.041
MQ54	5.000	2.576	1.158	0.356	0.612	0.613	0.566	0.035
MQ7	7.000	2.832	1.263	0.133	0.647	0.648	0.592	0.028
CC2-61	3.000	2.093	0.855	0.015	0.522	0.523	0.441	0.038
CC2-54	5.000	2.762	1.209	0.378	0.638	0.639	0.591	0.028
CC2-48	7.000	3.348	1.301	0.526	0.701	0.702	0.646	0.023
MQ59	7.000	2.752	1.294	0.016	0.637	0.638	0.590	0.034
MQ32	9.000	2.753	1.286	0.019	0.637	0.638	0.588	0.055
MQ37	7.000	1.391	0.556	0.099	0.281	0.282	0.257	0.051
MQ39	9.000	2.856	1.359	0.019	0.650	0.651	0.609	0.050
MQ47	7.000	2.107	0.953	0.012	0.525	0.526	0.451	0.033
MQ12	8.000	2.977	1.361	0.028	0.664	0.665	0.621	0.052
MQ64	6.000	2.379	1.016	0.016	0.580	0.581	0.500	0.035
MQ16	7.000	1.372	0.606	0.015	0.271	0.272	0.258	0.050
RPly02	10.000	3.062	1.468	0.557	0.673	0.674	0.642	0.026
RPly22	9.000	1.538	0.797	0.347	0.350	0.350	0.334	0.018
RPly109	23.000	6.675	2.334	0.690	0.850	0.852	0.838	0.055
平均值Mean	7.941	2.808	1.200	0.191	0.588	0.589	0.544	0.038

群体 Population	N_a	N_e	I	H_o	H_e	H
北京 Beijing	3.412	2.072	0.845	0.136	0.481	0.497
山西 Shanxi	4.765	2.772	1.136	0.164	0.582	0.595
山东 Shandong	6.412	2.897	1.175	0.194	0.588	0.590
美国阿巴拉契亚 Appalachia of USA	6.353	2.501	1.113	0.207	0.523	0.530
辽宁 Liaoning	4.882	2.865	1.177	0.208	0.609	0.621
河南 Henan	4.118	2.586	1.058	0.183	0.578	0.588
平均值Mean	4.990	2.616	1.084	0.182	0.560	0.570

变异来源 Source variation	方差总和 Sum of squares	平均方差 Mean of squares	变异分量 Estimated variance component	变异百分比 Percentage of variation (%)
群体间Among populations	71.360	14.272	0.077	2
个体间Among individuals	2 560.360	8.077	3.233	66
个体内Within individuals	520.500	1.611	1.611	32
总和Total	3 152.220		4.921	100

取样比例 Sampling ratio (%)	N_a	N_e	I	H_o	H_e	H	等位基因总数 Total allele
100	7.941	2.808	1.200	0.191	0.588	0.589	135
60	7.824	3.491	1.363	0.214	0.658	0.660	133
55	7.824	3.613	1.390	0.221	0.670	0.672	133
50	7.765	3.668	1.405	0.222	0.676	0.679	132
45	7.647	3.720	1.415	0.225	0.679	0.682	130
40	7.412	3.807	1.433	0.227	0.689	0.691	126
35	7.412	3.918	1.464	0.226	0.701	0.704	126
30	7.588	3.829	1.461	0.234	0.694	0.698	129
25	7.412	3.938	1.473	0.238	0.698	0.702	126
20	7.235	4.060	1.501	0.254	0.708	0.714	123
15	7.059	3.944	1.486	0.260	0.703	0.710	120
10	6.529	3.542	1.408	0.281	0.669	0.680	111
5	5.941	3.467	1.392	0.213	0.667	0.689	101