常绿杜鹃种质遗传多样性分析及SSR指纹图谱构建

doi:10.16420/j.issn.0513-353x.2024-0158

摘要/Abstract

摘要：

利用马缨杜鹃（Rhododendron delavayi Franch.）基因组测序数据进行genomic-SSR的分布特征分析和多态性SSR引物开发，并对61份常绿杜鹃种质进行亲缘关系分析和指纹图谱构建，以实现种质间亲缘关系聚类及其亲本来源推测。开发出了15对多态性高、稳定性好、亚属间通用性强的SSR引物，在61份种质中共检测到等位基因位点126个，每对引物的平均等位基因数（N_a）为8.400，平均有效等位基因数（N_e）为3.4681，平均观测杂合度（H_o）和期望杂合度（H_e）分别为0.3421和0.5347，平均多态信息含量（PIC）为0.6338。群体遗传结构分析结果将61份常绿杜鹃种质分成3个亚类群，与亲缘关系聚类结果基本一致，结合已知谱系资料推测所引进日本品种的祖先亲本来源于两个不同的分布地，且包括自育品种在内的所有品种均是以常绿杜鹃亚属（subgen. Hymenanthes）常绿杜鹃组（sect. Ponticum）内物种为亲本的杂交种。利用5个SSR标记成功构建出61份常绿杜鹃种质的指纹图谱。

关键词: 常绿杜鹃, 基因组SSR标记, 遗传多样性, 指纹图谱

Abstract:

In this study，based on genomic sequencing data of Rhododendron delavayi Franch.，genomic-SSR distribution characteristics were analyzed and polymorphic SSR primers were developed. Furthermore，the 61 evergreen Rhododendron germplasms were analyzed for genetic relationship and fingerprint construction to realize their genetic relationships and speculate their possible parents. In the results，15 pairs of SSR primers with high polymorphism，good stability and strong intergeneric versatility were developed，and a total of 126 alleles were detected in 61 evergreen Rhododendron germplasms. The average number of alleles（N_a）and the average effective number of alleles（N_e）were 8.400 and 3.4681，respectively. The mean observed heterozygosity（H_o）and expected heterozygosity（H_e）were 0.3421 and 0.5347，respectively，and the mean polymorphism information content（PIC）was 0.5163. According to the results of population genetic structure analysis，61 evergreen Rhododendron germplasms were divided into 3 subgroups，which were basically consistent with the results of phylogenetic analysis. Combined with the pedigree information，it was inferred that the ancestral parents of the introduced Japanese cultivars came from two different distribution areas，and all cultivars，including the self-bred cultivars，were the hybrids of the parental species from sect. Ponticum in subgen. Hymenanthes. Five SSR markers with high discriminability were used to construct the fingerprint of 61 evergreen Rhododendron germplasms successfully.

Key words: evergreen rhododendron, genomic SSR markers, genetic diversity, fingerprints

张露, 张平萍, 解玮佳, 许凤, 彭绿春, 宋杰, 杜光辉, 李世峰. 常绿杜鹃种质遗传多样性分析及SSR指纹图谱构建[J]. 园艺学报, 2025, 52(2): 380-394.

ZHANG Lu, ZHANG Pingping, XIE Weijia, XU Feng, PENG Lüchun, SONG Jie, DU Guanghui, LI Shifeng. Genetic Diversity Analysis and SSR Fingerprint Construction of Evergreen Rhododendron Germplasms[J]. Acta Horticulturae Sinica, 2025, 52(2): 380-394.

图/表 9

参考文献 36

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序号 No.	编号 ID	种/品种名 Species/cultivars name	分类/谱系（♀ × ♂） Classification/pedigree
1	WS-1	大喇叭杜鹃R. excellens	杜鹃亚属杜鹃组有鳞大花亚组subgen. Rhododendron sect. Rhododendron subsect. Maddenia
2	WS-3	露珠杜鹃R. irroratum	常绿杜鹃亚属常绿杜鹃组露珠杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect. Irrorata
3	WS-4	大白杜鹃R. decorum	常绿杜鹃亚属常绿杜鹃组云锦杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect. Fortunea
4	WS-5	滇南杜鹃R. hancockii	马银花亚属长蕊组subgen. Azaleastrum sect. Choniastrum
5	WS-6	云南杜鹃R. yunnanense	杜鹃亚属杜鹃组三花杜鹃亚组subgen. Rhododendron sect. Rhododendron subsect.Triflora
6	WS-7	宽杯杜鹃R. sinofalconeri	常绿杜鹃亚属常绿杜鹃组杯毛杜鹃亚组 subgen. Hymenanthes sect. Ponticum subsect. Falconera
7	WS-8	马缨杜鹃R. delavayi	常绿杜鹃亚属常绿杜鹃组树形杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect.Arborea
8	WS-9	马银花杜鹃R. ovatum	马银花亚属马银花组subgen. Azaleastrum sect. Azaleastrum
9	W6	凯特Catawbiense Grandiflorum	Catawbiense × 不详Unknown
10	W13	伊优克 Eucharitis	不详Unknown
11	W14	闪烁 Goldflimmer	catawbiense or Ponticum seedling
12	W17	莱姆 Halfdan Lem	The Hon. Jean Marie de Montague × Red Loderi
13	W19	门铃 Jingle Bells	Fabia group × Ole Olson
14	W21	罗伯茨 Lord Roberts	Catawbiense × 不详Unknown
15	W27	红杰克 Red Jack	Wilgen’s Ruby × May Day
16	W29	星光 Scintillation	不详Unknown
17	W31	玳瑁橙 Tortoiseshell Orange	Tortoiseshell group
18	W33	红珍珠 Wilgen’s Ruby	Britannia × John Walter
19	W35	锦缎 Germania	Antoon van Welie × Williamsianum
20	R1	诺娃 Nova Zembla	Parsons Grandiflorum × dark red hybrid
21	R2	粉黛 XXL	President Roosevelt × W35
22	J1	普罗米修斯 Prominence	不详Unknown
23	J2	紫色彩虹 Purple Rainbow	不详Unknown
24	J3	紫色宫殿 Purple Dome	不详Unknown
25	J4	红蝶 Red Monarch	不详Unknown
26	J5	薰衣草王子 Lavendor Prince	不详Unknown
27	J6	草莓圣代 Strawberry Sundae	不详Unknown
28	J7	珍珠姬 Sin Juhime	不详Unknown
29	J8	玛丽亚 Marian	不详Unknown
30	J9	甜心 Sweet Heart	不详Unknown
31	J10	阿特拉斯 Atlas	不详Unknown
32	J11	藤井夫人 Mrs Fujii	不详Unknown
33	J13	婚礼花束 Wedding Bouquet	不详Unknown
34	J14	太阳 Sun	不详Unknown
35	J16	彩虹 Rainbow	不详Unknown
36	J18	雨滴 Raindrop	不详Unknown
37	J19	火把 Torch	不详Unknown
38	J20	大地 Earth	不详Unknown
39	ZY-1	金色铃铛 Jinse Lingdang	W19 × 大白杜鹃 W19 × R. decorum
40	ZY-2	贵妃醉酒 Guifei Zuijiu	W13 × 大白杜鹃 W13 × R. decorum
41	ZY-3	云之绯羽 Yunzhifeiyu	W29 × 露珠杜鹃 W29 × R. irroratum
42	ZY-4	云之染彩 Yunzhirancai	R1 × 大白杜鹃 R1 × R. decorum
43	ZY-5	翠湖晓妆 Cuihu Xiaozhuang	W6 × R2
44	ZY-6	翠湖怜香 Cuihu Lianxiang	W21 × 大白杜鹃 W21 × R. decorum
45	ZY-7	红妆 Hongzhuang	W35 × R2
46	ZY-8	芳华 Fanghua	W35 × R2
47	ZY-9	黛玉 Daiyu	W35 × R2
48	ZY-10	花妍 Huayan	W35 × R2
49	ZY-11	墨碟心 Modiexin	（Ponticum × Unknown）
50	ZY-12	粉精灵 Fenjingling	大白杜鹃芽变 Bud sport of R. decorum
51	ZY-13	云之蜜语 Yunzhimiyu	W13 × 大白杜鹃 W13 × R. decorum
52	ZY-14	阳春雪 Yangchunxue	W35 × R2
53	ZY-15	宝钗 Baochai	大白杜鹃 × R2 R. decorum × R2
54	ZY-16	湘云 Xiangyun	R2 × 露珠杜鹃 R2 × R. irroratum
55	ZY-17	怡红 Yihong	R2 × 马缨杜鹃 R2 × R. delavayi
56	ZY-18	落雁 Luoyan	（Ponticum × Unknown）
57	ZY-19	昆上初红 Kunshang Chuhong	大白杜鹃 × 马缨杜鹃 R. decorum × R. delavayi
58	ZY-20	罗裳 Luochang	W29 × 马塞尔（Ponticum × Unknown）
59	ZY-21	翠湖紫鸥 Cuihu Ziou	安娜（Ponticum × Unknown）× R1
60	ZY-22	翠湖彤云 Cuihu Tongyun	安娜（Ponticum × Unknown）× R1
61	ZY-23	翠湖涟漪 Cuihu Lianyi	大白杜鹃 × W6 R. decorum × W6

序号 No.	编号 ID	种/品种名 Species/cultivars name	分类/谱系（♀ × ♂） Classification/pedigree
1	WS-1	大喇叭杜鹃R. excellens	杜鹃亚属杜鹃组有鳞大花亚组subgen. Rhododendron sect. Rhododendron subsect. Maddenia
2	WS-3	露珠杜鹃R. irroratum	常绿杜鹃亚属常绿杜鹃组露珠杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect. Irrorata
3	WS-4	大白杜鹃R. decorum	常绿杜鹃亚属常绿杜鹃组云锦杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect. Fortunea
4	WS-5	滇南杜鹃R. hancockii	马银花亚属长蕊组subgen. Azaleastrum sect. Choniastrum
5	WS-6	云南杜鹃R. yunnanense	杜鹃亚属杜鹃组三花杜鹃亚组subgen. Rhododendron sect. Rhododendron subsect.Triflora
6	WS-7	宽杯杜鹃R. sinofalconeri	常绿杜鹃亚属常绿杜鹃组杯毛杜鹃亚组 subgen. Hymenanthes sect. Ponticum subsect. Falconera
7	WS-8	马缨杜鹃R. delavayi	常绿杜鹃亚属常绿杜鹃组树形杜鹃亚组subgen. Hymenanthes sect. Ponticum subsect.Arborea
8	WS-9	马银花杜鹃R. ovatum	马银花亚属马银花组subgen. Azaleastrum sect. Azaleastrum
9	W6	凯特Catawbiense Grandiflorum	Catawbiense × 不详Unknown
10	W13	伊优克 Eucharitis	不详Unknown
11	W14	闪烁 Goldflimmer	catawbiense or Ponticum seedling
12	W17	莱姆 Halfdan Lem	The Hon. Jean Marie de Montague × Red Loderi
13	W19	门铃 Jingle Bells	Fabia group × Ole Olson
14	W21	罗伯茨 Lord Roberts	Catawbiense × 不详Unknown
15	W27	红杰克 Red Jack	Wilgen’s Ruby × May Day
16	W29	星光 Scintillation	不详Unknown
17	W31	玳瑁橙 Tortoiseshell Orange	Tortoiseshell group
18	W33	红珍珠 Wilgen’s Ruby	Britannia × John Walter
19	W35	锦缎 Germania	Antoon van Welie × Williamsianum
20	R1	诺娃 Nova Zembla	Parsons Grandiflorum × dark red hybrid
21	R2	粉黛 XXL	President Roosevelt × W35
22	J1	普罗米修斯 Prominence	不详Unknown
23	J2	紫色彩虹 Purple Rainbow	不详Unknown
24	J3	紫色宫殿 Purple Dome	不详Unknown
25	J4	红蝶 Red Monarch	不详Unknown
26	J5	薰衣草王子 Lavendor Prince	不详Unknown
27	J6	草莓圣代 Strawberry Sundae	不详Unknown
28	J7	珍珠姬 Sin Juhime	不详Unknown
29	J8	玛丽亚 Marian	不详Unknown
30	J9	甜心 Sweet Heart	不详Unknown
31	J10	阿特拉斯 Atlas	不详Unknown
32	J11	藤井夫人 Mrs Fujii	不详Unknown
33	J13	婚礼花束 Wedding Bouquet	不详Unknown
34	J14	太阳 Sun	不详Unknown
35	J16	彩虹 Rainbow	不详Unknown
36	J18	雨滴 Raindrop	不详Unknown
37	J19	火把 Torch	不详Unknown
38	J20	大地 Earth	不详Unknown
39	ZY-1	金色铃铛 Jinse Lingdang	W19 × 大白杜鹃 W19 × R. decorum
40	ZY-2	贵妃醉酒 Guifei Zuijiu	W13 × 大白杜鹃 W13 × R. decorum
41	ZY-3	云之绯羽 Yunzhifeiyu	W29 × 露珠杜鹃 W29 × R. irroratum
42	ZY-4	云之染彩 Yunzhirancai	R1 × 大白杜鹃 R1 × R. decorum
43	ZY-5	翠湖晓妆 Cuihu Xiaozhuang	W6 × R2
44	ZY-6	翠湖怜香 Cuihu Lianxiang	W21 × 大白杜鹃 W21 × R. decorum
45	ZY-7	红妆 Hongzhuang	W35 × R2
46	ZY-8	芳华 Fanghua	W35 × R2
47	ZY-9	黛玉 Daiyu	W35 × R2
48	ZY-10	花妍 Huayan	W35 × R2
49	ZY-11	墨碟心 Modiexin	（Ponticum × Unknown）
50	ZY-12	粉精灵 Fenjingling	大白杜鹃芽变 Bud sport of R. decorum
51	ZY-13	云之蜜语 Yunzhimiyu	W13 × 大白杜鹃 W13 × R. decorum
52	ZY-14	阳春雪 Yangchunxue	W35 × R2
53	ZY-15	宝钗 Baochai	大白杜鹃 × R2 R. decorum × R2
54	ZY-16	湘云 Xiangyun	R2 × 露珠杜鹃 R2 × R. irroratum
55	ZY-17	怡红 Yihong	R2 × 马缨杜鹃 R2 × R. delavayi
56	ZY-18	落雁 Luoyan	（Ponticum × Unknown）
57	ZY-19	昆上初红 Kunshang Chuhong	大白杜鹃 × 马缨杜鹃 R. decorum × R. delavayi
58	ZY-20	罗裳 Luochang	W29 × 马塞尔（Ponticum × Unknown）
59	ZY-21	翠湖紫鸥 Cuihu Ziou	安娜（Ponticum × Unknown）× R1
60	ZY-22	翠湖彤云 Cuihu Tongyun	安娜（Ponticum × Unknown）× R1
61	ZY-23	翠湖涟漪 Cuihu Lianyi	大白杜鹃 × W6 R. decorum × W6

SSR类型 SSR type	类型数 Number	重复次数 Repeats	占比/% Ratios	重复基序 Repeat motifs	SSR数量 Number of SSR	总占比/% Total ratios
单核苷酸 Mono-nucleotide	2	≥ 10	35.19	A/T C/G	41 699 913	34.01 0.74
二核苷酸 Di-nucleotide	6	≥ 6	41.63	AA/TT AC/GT AG/CT AT/AT CC/GG CG/CG	16 900 4 444 24 151 4 448 361 109	13.78 3.62 19.70 3.63 0.29 0.09
三核苷酸 Tri-nucleotide	12	≥ 5	10.23	AAA/TTT AAC/GTT AAG/CTT AAT/ATT ACC/GGT 其他 Others	5 260 582 1 786 630 1 523 4 131	4.29 0.47 1.46 0.51 1.24 3.37
四核苷酸 Tetra-nuleotide	33	≥ 5	9.47	AAAA/TTTT AAAC/GTTT AAAG/CTTT AAAT/ATTT 其他 Others	368 36 77 159 10 828	0.30 0.03 0.06 0.13 8.83
五核苷酸 Penta-nucleotide	39	≥ 5	0.20	AAAAG/CTTTT 其他 Others	48 203	0.04 0.17
六核苷酸 Hexa-nucleotide	85	≥ 5	3.28	AAAAAG/CTTTTT 其他 Others	13 3 956	0.01 3.23
合计Total	177		100.00		122 625	100.00

SSR类型 SSR type	类型数 Number	重复次数 Repeats	占比/% Ratios	重复基序 Repeat motifs	SSR数量 Number of SSR	总占比/% Total ratios
单核苷酸 Mono-nucleotide	2	≥ 10	35.19	A/T C/G	41 699 913	34.01 0.74
二核苷酸 Di-nucleotide	6	≥ 6	41.63	AA/TT AC/GT AG/CT AT/AT CC/GG CG/CG	16 900 4 444 24 151 4 448 361 109	13.78 3.62 19.70 3.63 0.29 0.09
三核苷酸 Tri-nucleotide	12	≥ 5	10.23	AAA/TTT AAC/GTT AAG/CTT AAT/ATT ACC/GGT 其他 Others	5 260 582 1 786 630 1 523 4 131	4.29 0.47 1.46 0.51 1.24 3.37
四核苷酸 Tetra-nuleotide	33	≥ 5	9.47	AAAA/TTTT AAAC/GTTT AAAG/CTTT AAAT/ATTT 其他 Others	368 36 77 159 10 828	0.30 0.03 0.06 0.13 8.83
五核苷酸 Penta-nucleotide	39	≥ 5	0.20	AAAAG/CTTTT 其他 Others	48 203	0.04 0.17
六核苷酸 Hexa-nucleotide	85	≥ 5	3.28	AAAAAG/CTTTTT 其他 Others	13 3 956	0.01 3.23
合计Total	177		100.00		122 625	100.00

编号 Code	位点 Loci	染色体号 Chr.	重复单元 Repeat unit	荧光标记 Fluorescent dye	引物序列（5′-3′） Primer Sequences	产物/bp Product	PIC
RdSSR01	Rhdel01G0240300	1	(AAA)₅	FAM	F：CTCTCAAATATTCGCCAGCC； R：CCTCTTCTGTCGGGTGAAAC	113 ~ 116	0.6132
RdSSR02	Rhdel03G0011900	3	(CGG)₆	FAM	F：ACTGGCGTTTGGAATTTGAC； R：TCCAAAGCAATCTTCCATCC	199 ~ 221	0.4730
RdSSR03	Rhdel03G0235100	3	(CGG)₅	HEX	F：CACGAGATTGGGAGTACGGT； R：AGCTGCGAGTATCCCAGTGT	278 ~ 287	0.8942
RdSSR04	Rhdel03G0250000	3	(AAG)₅	HEX	F：GCGTTACCGGCAGTATTGTT； R：GTGGCCTCTAGCCTTGTGAG	207 ~ 241	0.6773
RdSSR05	Rhdel03G0272700	3	(CTT)₅	FAM	F：CAACCAAGCAAGTGCAAAGA； R：CTGAGGCCTGTCTGGAAGTC	235 ~ 257	0.6111
RdSSR06	Rhdel04G0032400	4	(GCA)₇	HEX	F：GAATTCTTTCGCCACTCTGC； R：AAGCCATGGAGTTGATACGG	128 ~ 144	0.7279
RdSSR07	Rhdel04G0036900	4	(CCG)₆	HEX	F：ACGCCGCTAATAACCACAAC； R：GATAAATCCGGTGAATCCGA	224 ~ 240	0.5445
RdSSR08	Rhdel04G0139400	4	(GA)₈	HEX	F：CTCACGTGGTCCATCATTTG； R：TAGGGTTCCATCACAAAGCC	291 ~ 307	0.8265
RdSSR09	Rhdel04G0358500	4	(TC)₉	HEX	F：TGTGGAACATCTTGAGCAGC； R：AGTCAACAGGCCAAACCAAC	127 ~ 204	0.7858
RdSSR10	Rhdel05G0013300	5	(AG)₉	HEX	F：GGTGCACCAAGGTCTTCATT； R：ATCTATACCCGCTGCAATCG	281 ~ 288	0.6934
RdSSR11	Rhdel05G0137700	5	(GGA)₆	HEX	F：CATGCTGAAGAGTTGGACGA； R：CTTTGGAGACTGGAGGCTTG	151 ~ 155	0.2392
RdSSR12	Rhdel08G0018000	8	(AG)₈	FAM	F：GCTATGGAGTGCTTTAGCCG； R：TGATTGCTACGATCGCTTTG	131 ~ 150	0.4846
RdSSR13	Rhdel12G0119900	12	(AAC)₉	TAM	F：TCATCGACATCGTCAGCTTC； R：GCCTGATTTCACCTCGTGAT	227 ~ 245	0.7038
RdSSR14	Rhdel12G0190300	12	(TTC)₇	FAM	F：GGAGCAGCAGAACAATGAC； R：GTAAAGGGCGTTCGTATGGA	189 ~ 339	0.8926
RdSSR15	Rhdel13G0101600	13	(TC)₈	FAM	F：CCATGGAAATTGATTGGACC； R：ACAAATGGGCTAACTGGACG	184 ~ 296	0.3402
平均 Average							0.6338