结合分子手段和形态分析验证大白杜鹃与马缨杜鹃的自然杂交

doi:10.17520/biods.2017090

生物多样性 ›› 2017, Vol. 25 ›› Issue (6): 627-637. DOI: 10.17520/biods.2017090

结合分子手段和形态分析验证大白杜鹃与马缨杜鹃的自然杂交

郑硕理^1,², 田晓玲³, 黄承玲³, 王灵军⁴, 冯元⁴, 张敬丽^1,^*()

1 云南农业大学园林园艺学院, 昆明 650201
2 湖南省森林植物园, 长沙 410116
3 贵州民族大学人文科技学院, 贵阳 550025
4 贵州百里杜鹃科研所, 贵州毕节 551518

收稿日期:2017-03-22 接受日期:2017-06-28 出版日期:2017-06-20 发布日期:2017-07-10
通讯作者: 张敬丽
基金资助:
国家自然科学基金(31260079)和国家科技支撑计划(2015BAD10B00)

Molecular and morphological evidence for natural hybridization between Rhododendron decorum and R. delavayi (Ericaceae)

Shuoli Zheng^1,², Xiaoling Tian³, Chengling Huang³, Lingjun Wang⁴, Yuan Feng⁴, Jingli Zhang^1,^*()

1 College of Landscape and Gardening, Yunnan Agriculture University, Kunming 650201
2 Hunan Forest Botanical Garden, Changsha 410116
3 College of Humanities & Science, Guizhou Minzu University, Guiyang 550025
4 Institute of Guizhou Baili Rhododendron, Bijie, Guizhou 551518

Received:2017-03-22 Accepted:2017-06-28 Online:2017-06-20 Published:2017-07-10
Contact: Zhang Jingli

摘要/Abstract

摘要：

中国西南地区是世界杜鹃属(Rhododendron)植物的分布和分化中心, 开展杜鹃属自然杂交研究可为分类修订和新品种选育提供依据。本文以贵州百里杜鹃自然保护区(百里保护区)和云南石宝山分布的大白杜鹃(Rhododendron decorum)、马缨杜鹃(R. delavayi)及两种间疑似自然杂交后代和人工杂交后代个体为研究材料, 通过形态特征比较、核基因间隔序列ITS以及叶绿体基因片段trnL-trnF、rbcL和trnH-psbA测序特征分析, 发现疑似自然杂交后代与人工杂交后代形态特征介于大白杜鹃与马缨杜鹃之间; 4个DNA片段序列均能区分大白杜鹃和马缨杜鹃, 同一物种的不同居群在叶绿体基因片段rbcL、trnH-psbA序列存在差异位点; 所有人工杂交后代个体和1个石宝山疑似杂交个体均以大白杜鹃为母本, 所有百里保护区疑似杂交个体和6个石宝山疑似杂交个体以马缨杜鹃为母本。本研究证实了大白杜鹃与马缨杜鹃在百里保护区和云南石宝山存在自然杂交现象, 杂交为双向且自然杂交后代绝大多数以马缨杜鹃为母本。

关键词: 杜鹃属, 自然杂交, 形态特征, DNA序列

Abstract

The southwestern region of China is the center of differentiation for the genus Rhododendron. Studies on the natural hybridization of Rhododendron provide solutions for taxonomic problems and enhance the ability to breed new cultivars faster. In this paper, we compared the morphological features and sequenced nuclear internal transcribed spacer (ITS) and chloroplast trnL-trnF, rbcL, trnH-psbA for Rhododendron decorum, R. delavayi, and the putative hybrid offspring of these two species. All samples in this study were collected from Baili Rhododendron Nature Reserve in Guizhou Province, the Shibao Mountain of Yunnan Province and Kunming Botanical Garden. Results showed that all putative hybrids (natural and artificial) produced an intermediate morphology. Sequencing results revealed that all artificial hybrids and one collected from the Shibao Mountain inherited cpDNA from R. decorum, and all putative hybrids collected from Baili Rhododendron Nature Reserve and six from Shibao Mountain inherited cpDNA from R. delavayi. In this study, we have confirmed that natural hybridization between R. decorum and R. delavayi exists in the Guizhou and Yunnan sites. Hybridization was bilateral, and most of the hybrids were derived from maternal parent R. delavayi.

Key words: Rhododendron, natural hybridization, morphological traits, DNA sequences

郑硕理, 田晓玲, 黄承玲, 王灵军, 冯元, 张敬丽 (2017) 结合分子手段和形态分析验证大白杜鹃与马缨杜鹃的自然杂交. 生物多样性, 25, 627-637. DOI: 10.17520/biods.2017090.

Shuoli Zheng, Xiaoling Tian, Chengling Huang, Lingjun Wang, Yuan Feng, Jingli Zhang (2017) Molecular and morphological evidence for natural hybridization between Rhododendron decorum and R. delavayi (Ericaceae). Biodiversity Science, 25, 627-637. DOI: 10.17520/biods.2017090.

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https://www.biodiversity-science.net/CN/Y2017/V25/I6/627

图/表 9

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编号 Code	种类 Taxon	地点 Site	经纬度 Latitude and longitude	海拔 Altitude (m)	个体数量 Sample size
BLM (1-10)	马缨杜鹃 R. delavayi	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	10
BLD (1-10)	大白杜鹃 R. decorum	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	10
BLH (1-8)	疑似杂交后代 Putative hybrids	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	8
SBSM (1-5)	马缨杜鹃 R. delavayi	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	5
SBSD (1-5)	大白杜鹃 R. decorum	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	5
SBSH (2/3/4/5/7/8/9)	疑似杂交后代 Putative hybrids	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	7
AH (1-5)	人工杂交后代 Artificial hybrids	昆明植物园 Kunming Botanical Garden			5
总计 Total					50

编号 Code	种类 Taxon	地点 Site	经纬度 Latitude and longitude	海拔 Altitude (m)	个体数量 Sample size
BLM (1-10)	马缨杜鹃 R. delavayi	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	10
BLD (1-10)	大白杜鹃 R. decorum	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	10
BLH (1-8)	疑似杂交后代 Putative hybrids	贵州百里杜鹃自然保护区 Baili Rhododendron Nature Reserve in Guizhou Province	27°15′ N 105°58′ E	1,700-1,800	8
SBSM (1-5)	马缨杜鹃 R. delavayi	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	5
SBSD (1-5)	大白杜鹃 R. decorum	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	5
SBSH (2/3/4/5/7/8/9)	疑似杂交后代 Putative hybrids	云南大理石宝山 Shibao Mountain of Dali, Yunnan	26°22′ N 99°50′ E	2,400-2,600	7
AH (1-5)	人工杂交后代 Artificial hybrids	昆明植物园 Kunming Botanical Garden			5
总计 Total					50

形态特征 Morphological characters	大白杜鹃 R. decorum	疑似自然杂交与人工杂交后代 Putative and artificial hybrids	马缨杜鹃 R. delavayi
花冠颜色 Corolla color	白色 White	粉红 Pink	红色 Red
花冠裂片数 No. of corolla lobes	6-8, 通常7 6-8, usually 7	5-6, 无规律 5-6, irregular	5
叶片形状 Leaf shape	长圆形 Oblong	长圆形至披针形 Oblong to lanceolate	披针形 Lanceolate
叶片毛被 Ventral leaf suraface indumentum	无毛 Glabrous	稀疏毛被 Thin indumentum	浓密海绵状毛被 Woolly indumentum
果实表面特征 Fruit surface character	有腺体 Glands	有腺体或毛被或两者均有 Glands or/and indumentum	密被棕红色毛被 Continuous brownish red indumentum
花期 Flowering period	5-6月 May to June	4月中旬-5月中旬 Mid-April to mid-May	3月-5月 March to May

形态特征 Morphological characters	大白杜鹃 R. decorum	疑似自然杂交与人工杂交后代 Putative and artificial hybrids	马缨杜鹃 R. delavayi
花冠颜色 Corolla color	白色 White	粉红 Pink	红色 Red
花冠裂片数 No. of corolla lobes	6-8, 通常7 6-8, usually 7	5-6, 无规律 5-6, irregular	5
叶片形状 Leaf shape	长圆形 Oblong	长圆形至披针形 Oblong to lanceolate	披针形 Lanceolate
叶片毛被 Ventral leaf suraface indumentum	无毛 Glabrous	稀疏毛被 Thin indumentum	浓密海绵状毛被 Woolly indumentum
果实表面特征 Fruit surface character	有腺体 Glands	有腺体或毛被或两者均有 Glands or/and indumentum	密被棕红色毛被 Continuous brownish red indumentum
花期 Flowering period	5-6月 May to June	4月中旬-5月中旬 Mid-April to mid-May	3月-5月 March to May

编号 Code	克隆数量 Clone numbers	类型 Type	差异位点 Variable sites (bp)						类型 Type
编号 Code	克隆数量 Clone numbers	类型 Type	94	101	114	203	494	505	A1	A2	A4
大白杜鹃 R. decorum	0	A1	T	G	G	T	T	C	15	0	0
马缨杜鹃 R. delavayi	0	A2	C	C	T	G	C	T	0	15	0
AH、BLH、SBSH		A3	Y	S	K	K	Y	Y
克隆测序 Clone sequences
BLH1	2								1	1	0
BLH2	4								1	2	1
BLH3	3								2	1	0
BLH4	3								2	1	0
BLH5	3								1	1	1
BLH6	5								1	1	3
BLH7	4								2	1	1
BLH8	4								2	1	1
AH1	3								1	2	1
AH2	3								1	2	0
AH3	4								2	2	0
AH4	3								2	1	0
AH5	4								2	2	1
SBSH2	6								4	2	0
SBSH3	8								5	3	0
SBSH4	5								4	1	0
SBSH5	5								4	1	0
SBSH7	7								5	2	0
SBSH8	7								6	1	0
SBSH9	5								3	2	0
克隆数量总计 Total clone numbers									53	31	10

结合分子手段和形态分析验证大白杜鹃与马缨杜鹃的自然杂交

Molecular and morphological evidence for natural hybridization between Rhododendron decorum and R. delavayi (Ericaceae)

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编号 Code	类型 Type	差异位点 Variable sites (bp)
编号 Code	类型 Type	147	622	623	624	626	627	629	646	831
大白杜鹃 Rhododendron decorum	B1	G	T	T	T	T	T	T	C	T
马缨杜鹃 R. delavayi	B2	T	A	A	A	A	A	A	A	C
AH、SBSH5	B1	G	T	T	T	T	T	T	C	T
BLH、SHSH2/3/4/7/8/9	B2	T	A	A	A	A	A	A	A	C

编号 Code	类型 Type	差异位点 Variable sites (bp)
编号 Code	类型 Type	8	30	263	398	407
BLD	C1	C	G	T	C	C
BLM	C2	C	T	A	T	T
SBSD	C3	G	G	T	C	C
SBSM	C4	G	T	A	T	T
AH	C1	C	G	T	C	C
BLH	C2	C	T	A	T	T
SBSH5	C3	G	G	T	C	C
SBSH2/3/4/7/8/9	C4	G	T	A	T	T

编号 Code	类型 Type	差异位点 Variable sites (bp)
编号 Code	类型 Type	1	2	128	135	211	214	245	254	254	299	313	333	337	340	344	348	351	358	365-369
BLD	D1	G	A	T	C	G	C	G	A	C	A	A	G	T	G	G	T	T	T	AATTC
SBSD	D2	-	G	*	*	*	*	*	*	*	*	*	A	A	A	A	A	G	G	*****
BLM	D3	*	*	A	G	A	C	T	T	T	T	G	*	*	*	*	*	*	*	G----
SBSM	D4	*	*	A	G	A	A	T	T	T	T	G	*	*	*	*	*	*	*	G----
AH	D1	G	A	T	C	G	C	G	A	C	A	A	G	T	G	G	T	T	T	AATTC
SBSH5	D2	*	G	*	*	*	*	*	*	*	*	*	A	A	A	A	A	G	G	*****
BLH	D3	*	*	A	G	A	C	T	T	T	T	G	*	*	*	*	*	*	*	G----
SBSH2/3/4/7/8/9	D4	*	*	A	G	A	A	T	T	T	T	G	*	*	*	*	*	*	·	G----

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