中国石斛属植物遗传资源的AFLP分析

doi:10.16420/j.issn.0513-353x.2021-0625

摘要/Abstract

摘要：

中国是世界石斛属（Dendrobium）植物分布中心,其主要分布在云贵川等地区。采用扩增片段长度多态性（AFLP）分子标记方法,研究了石斛属92个种质的遗传多样性和亲缘关系。从24对引物组合中筛选出10对引物组合进行扩增,共扩增出2 329个位点,其中多态位点2 307个,占总扩增位点的99.04%。聚类分析将92个石斛种质,分为4个类群,种间遗传分化显著。AFLP分子标记将菱唇石斛和西畴石斛划为石斛组、金耳石斛划为寡花组,且研究结果表明,茎的形态和质地为石斛分类的重要性状,花的唇瓣和茎节的膨大非石斛分类的主要因素。

关键词: 兰科, 石斛属, AFLP, 扩增, 亲缘关系, 遗传多样性

Abstract:

Dendrobium plants are mainly distributed in Yunnan,Guizhou and Sichuan Province in China. China is the distribution center of Dendrobium plants in the world. In this study,Amplified Length Polymorphism（AFLP）was used to study the genetic diversity and phylogenetic relationship of 92 species of Dendrobium in China. Ten primer combinations were screened out from 24 pairs of primer combinations,and a total of 2 329 loci were amplified,among which 2 307 were polymorphic,accounting for 99.04% of the total loci. Phylogenetic analysis results showed that 92 Dendrobium samples were divided into four groups,and the genetic differentiation was significant. AFLP molecular markers supported that the two species of D. leptocladum and D. xichouense were classified into Sect. Dendrobium,and D. hookerianum was divided into Sect. Holochrysa. The results also showed that the shape and texture of the stem were important characteristics for the classification of Dendrobium,and the flower lip and enlargement of stem node were not the main factors for Dendrobium classification.

Key words: Orchidaceae, Dendrobium, AFLP, amplification, phylogenetic relationship, genetic diversity

中图分类号:

S682.31

张萌, 单玉莹, 杨业波, 翟飞飞, 王兆山, 巨关升, 孙振元, 李振坚. 中国石斛属植物遗传资源的AFLP分析[J]. 园艺学报, 2022, 49(6): 1339-1350.

ZHANG Meng, SHAN Yuying, YANG Yebo, ZHAI Feifei, WANG Zhaoshan, JU Guansheng, SUN Zhenyuan, LI Zhenjian. AFLP Analysis of Genetic Resources of Dendrobium from China[J]. Acta Horticulturae Sinica, 2022, 49(6): 1339-1350.

图/表 4

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引种地 Introducing site	种质 Species（cultivars）	引种地 Introducing site	种质 Species（cultivars）
云南景洪 Jinghong,Yunnan	长苏石斛 D. brymerianum	云南文山 Wenshan,Yunnan	翅梗石斛 D. trigonopus
	短棒石斛 D. capillipes		西畴石斛 D. xichouense
	草石斛 D. compactum	云南西双版纳 Xishuangbanna,Yunnan	黑毛石斛 D. williamsonii
	齿瓣石斛 D. devonianum	贵州兴义 Xingyi,Guizhou	钩状石斛 D. aduncum
	景洪石斛 D. exile		束花石斛 D. chrysanthum
	流苏石斛 D. fimbriatum		玫瑰石斛 D. crepidatum
	棒节石斛 D. findlayanum		叠鞘石斛 D. denneanum
	曲轴石斛 D. gibsonii		重唇石斛 D. hercoglossum
	杯鞘石斛 D. gratiosissimum		聚石斛 D. lindleyi
	苏瓣石斛 D. harveyanum		罗河石斛 D. lohohense
	疏花石斛 D. henryi		杓唇石斛 D. moschatum
	尖刀唇石斛 D. heterocarpum	贵州赤水 Chishui,Guizhou	金钗石斛 D. nobile
	金耳石斛 D. hookerianum	贵州铜仁 Tongren,Guizhou	梵净山石斛 D. fanjingshanense
	小黄花石斛 D. jenkinsii	广东始兴 Shixing,Guangdong	始兴石斛 D. shixingense
	长距石斛 D. longicornu	广西崇左 Chongzuo,Guangxi	檀香石斛 D. anosmum
	铁皮石斛 D. officinale	广西金秀 Jinxiu,Guangxi	矩唇石斛 D. linawianum
	少花石斛 D. parciflorum	广西凌云 Lingyun,Guangxi	美花石斛 D. loddigesii
	肿节石斛 D. pendulum	广西那坡 Napo,Guangxi	藏南石斛 D. monticola
	单葶草石斛 D. porphyrochilum	广西玉林 Yulin,Guangxi	密花石斛 D. densiflorum
	报春石斛 D. primulinum		滇桂石斛 D. guangxiense
	梳唇石斛 D. strongylanthum	安徽霍山 Huoshan,Anhui	霍山石斛 D. huoshanense
	叉唇石斛 D. stuposum	海南儋州 Danzhou,Hainan	竹枝石斛 D. salaccense
	广东石斛 D. wilsonii	海南广坝 Guangba,Hannan	广坝石斛 D. lagarum
	球花石斛 D. thyrsiflorum	海南琼中 Qiongzhong,Hainan	海南石斛 D. hainanense
	黄花石斛 D. dixanthum		华石斛 D. sinense
云南丽江 Lijiang,Yunnan	王亮石斛 D. wangliangii	河南南阳 Nanyang,Henan	曲茎石斛 D. flexicaule
云南勐腊 Mengla,Yunnan	勐腊石斛 D. chrysocrepis	河南南召 Nanzhao,Henan	河南石斛 D. henanense
	反瓣石斛 D. ellipsophyllum	四川夹江 Jiajiang,Sichuan	夹江石斛 D. jiajiangense
	针叶石斛 D. pseudotenellum	四川泸定 Luding,Sichuan	细叶石斛 D. hancockii
	勐海石斛 D. sinominutiflorum	台湾兰屿 Lanyu,Taiwan,China	燕石斛 D. equitans
	刀叶石斛 D. terminale		双花石斛 D. furcatopedicellatum
	大苞鞘石斛 D. wardianum		红花石斛 D. goldschmidtianum
	具槽石斛 D. sulcatum		吕宋石斛 D. luzonense
	喇叭唇石斛 D. lituiflorum		木石斛 D. crumenatum
	剑叶石斛 D. acinaciform	台湾台东Taidong,Taiwan,China	菱唇石斛 D. leptocladum
云南屏边 Pingbian,Yunnan	高山石斛 D. infundibulum	菲律宾 Philippines	紫晶石斛 D. amethystoglossum
	矮石斛 D. bellatulum		澳洲石斛 D. kingianum
	翅萼石斛 D. cariniferum	澳大利亚 Australia	黄喉石斛 D. signatum
	河口石斛 D. hekouense		宝石石斛 D. smillieae
云南文山 Wenshan,Yunnan	串珠石斛 D. falconeri	越南 Vietnam	黄玉石斛 D. topaziacum
	兜唇石斛 D. aphyllum	云南昆明 Kunming,Yunnan	红星 D.‘Akane’
	喉红石斛 D. christyanum		梦幻 D.‘Dream’
	线叶石斛 D. aurantiacum		火鸟 D.‘Fire Bird’
	鼓槌石斛 D. chrysotoxum		女王 D.‘Princess’
	晶帽石斛 D. crystallinum	海南三亚 Sanya,Hainan	树兰 Epidendrum secundum
	细茎石斛 D. moniliforme	海南昌江 Changjiang,Hainan	拟石斛 Oxystophyllum changjiangense
	紫瓣石斛 D. parishii	台湾台南 Tainan,Taiwan,China	武夷兰 Angraecum leonis
	紫婉石斛 D. transparens

引种地 Introducing site	种质 Species（cultivars）	引种地 Introducing site	种质 Species（cultivars）
云南景洪 Jinghong,Yunnan	长苏石斛 D. brymerianum	云南文山 Wenshan,Yunnan	翅梗石斛 D. trigonopus
	短棒石斛 D. capillipes		西畴石斛 D. xichouense
	草石斛 D. compactum	云南西双版纳 Xishuangbanna,Yunnan	黑毛石斛 D. williamsonii
	齿瓣石斛 D. devonianum	贵州兴义 Xingyi,Guizhou	钩状石斛 D. aduncum
	景洪石斛 D. exile		束花石斛 D. chrysanthum
	流苏石斛 D. fimbriatum		玫瑰石斛 D. crepidatum
	棒节石斛 D. findlayanum		叠鞘石斛 D. denneanum
	曲轴石斛 D. gibsonii		重唇石斛 D. hercoglossum
	杯鞘石斛 D. gratiosissimum		聚石斛 D. lindleyi
	苏瓣石斛 D. harveyanum		罗河石斛 D. lohohense
	疏花石斛 D. henryi		杓唇石斛 D. moschatum
	尖刀唇石斛 D. heterocarpum	贵州赤水 Chishui,Guizhou	金钗石斛 D. nobile
	金耳石斛 D. hookerianum	贵州铜仁 Tongren,Guizhou	梵净山石斛 D. fanjingshanense
	小黄花石斛 D. jenkinsii	广东始兴 Shixing,Guangdong	始兴石斛 D. shixingense
	长距石斛 D. longicornu	广西崇左 Chongzuo,Guangxi	檀香石斛 D. anosmum
	铁皮石斛 D. officinale	广西金秀 Jinxiu,Guangxi	矩唇石斛 D. linawianum
	少花石斛 D. parciflorum	广西凌云 Lingyun,Guangxi	美花石斛 D. loddigesii
	肿节石斛 D. pendulum	广西那坡 Napo,Guangxi	藏南石斛 D. monticola
	单葶草石斛 D. porphyrochilum	广西玉林 Yulin,Guangxi	密花石斛 D. densiflorum
	报春石斛 D. primulinum		滇桂石斛 D. guangxiense
	梳唇石斛 D. strongylanthum	安徽霍山 Huoshan,Anhui	霍山石斛 D. huoshanense
	叉唇石斛 D. stuposum	海南儋州 Danzhou,Hainan	竹枝石斛 D. salaccense
	广东石斛 D. wilsonii	海南广坝 Guangba,Hannan	广坝石斛 D. lagarum
	球花石斛 D. thyrsiflorum	海南琼中 Qiongzhong,Hainan	海南石斛 D. hainanense
	黄花石斛 D. dixanthum		华石斛 D. sinense
云南丽江 Lijiang,Yunnan	王亮石斛 D. wangliangii	河南南阳 Nanyang,Henan	曲茎石斛 D. flexicaule
云南勐腊 Mengla,Yunnan	勐腊石斛 D. chrysocrepis	河南南召 Nanzhao,Henan	河南石斛 D. henanense
	反瓣石斛 D. ellipsophyllum	四川夹江 Jiajiang,Sichuan	夹江石斛 D. jiajiangense
	针叶石斛 D. pseudotenellum	四川泸定 Luding,Sichuan	细叶石斛 D. hancockii
	勐海石斛 D. sinominutiflorum	台湾兰屿 Lanyu,Taiwan,China	燕石斛 D. equitans
	刀叶石斛 D. terminale		双花石斛 D. furcatopedicellatum
	大苞鞘石斛 D. wardianum		红花石斛 D. goldschmidtianum
	具槽石斛 D. sulcatum		吕宋石斛 D. luzonense
	喇叭唇石斛 D. lituiflorum		木石斛 D. crumenatum
	剑叶石斛 D. acinaciform	台湾台东Taidong,Taiwan,China	菱唇石斛 D. leptocladum
云南屏边 Pingbian,Yunnan	高山石斛 D. infundibulum	菲律宾 Philippines	紫晶石斛 D. amethystoglossum
	矮石斛 D. bellatulum		澳洲石斛 D. kingianum
	翅萼石斛 D. cariniferum	澳大利亚 Australia	黄喉石斛 D. signatum
	河口石斛 D. hekouense		宝石石斛 D. smillieae
云南文山 Wenshan,Yunnan	串珠石斛 D. falconeri	越南 Vietnam	黄玉石斛 D. topaziacum
	兜唇石斛 D. aphyllum	云南昆明 Kunming,Yunnan	红星 D.‘Akane’
	喉红石斛 D. christyanum		梦幻 D.‘Dream’
	线叶石斛 D. aurantiacum		火鸟 D.‘Fire Bird’
	鼓槌石斛 D. chrysotoxum		女王 D.‘Princess’
	晶帽石斛 D. crystallinum	海南三亚 Sanya,Hainan	树兰 Epidendrum secundum
	细茎石斛 D. moniliforme	海南昌江 Changjiang,Hainan	拟石斛 Oxystophyllum changjiangense
	紫瓣石斛 D. parishii	台湾台南 Tainan,Taiwan,China	武夷兰 Angraecum leonis
	紫婉石斛 D. transparens

接头 Adaptor	引物 Primer	序列（5′-3′） Sequence	引物 Primer	序列（5′-3′） Sequence
EcoR I-1		CTCGTAGACTGCGTACC	E83	GACTGCGTACCAATTCTCA
EcoR I-2		AATTGGTACGCAGTCTAC	M48	GATGAGTCCTGAGTAACAC
Mse I-1		GACGATGAGTCCTGAG	M49	GATGAGTCCTGAGTAACAG
Mse I-2		TACTCAGGACTCAT	M50	GATGAGTCCTGAGTAACAT
	EOO	GACTGCGTACCAATTC	M51	GATGAGTCCTGAGTAACCA
	MOO	GATGAGTCCTGAGTAA	M55	GATGAGTCCTGAGTAACGA
	E32	GACTGCGTACCAATTCAAC	M59	GATGAGTCCTGAGTAACTA
	E33	GACTGCGTACCAATTCAAG	M60	GATGAGTCCTGAGTAACTC
	E35	GACTGCGTACCAATTCACA	M61	GATGAGTCCTGAGTAACTG
	E37	GACTGCGTACCAATTCACG	M62	GATGAGTCCTGAGTAACTT
	E38	GACTGCGTACCAATTCACT	M84	GATGAGTCCTGAGTAATCC
	E75	GACTGCGTACCAATTCGTA

接头 Adaptor	引物 Primer	序列（5′-3′） Sequence	引物 Primer	序列（5′-3′） Sequence
EcoR I-1		CTCGTAGACTGCGTACC	E83	GACTGCGTACCAATTCTCA
EcoR I-2		AATTGGTACGCAGTCTAC	M48	GATGAGTCCTGAGTAACAC
Mse I-1		GACGATGAGTCCTGAG	M49	GATGAGTCCTGAGTAACAG
Mse I-2		TACTCAGGACTCAT	M50	GATGAGTCCTGAGTAACAT
	EOO	GACTGCGTACCAATTC	M51	GATGAGTCCTGAGTAACCA
	MOO	GATGAGTCCTGAGTAA	M55	GATGAGTCCTGAGTAACGA
	E32	GACTGCGTACCAATTCAAC	M59	GATGAGTCCTGAGTAACTA
	E33	GACTGCGTACCAATTCAAG	M60	GATGAGTCCTGAGTAACTC
	E35	GACTGCGTACCAATTCACA	M61	GATGAGTCCTGAGTAACTG
	E37	GACTGCGTACCAATTCACG	M62	GATGAGTCCTGAGTAACTT
	E38	GACTGCGTACCAATTCACT	M84	GATGAGTCCTGAGTAATCC
	E75	GACTGCGTACCAATTCGTA

引物 Primer	N_a	N_e	H	I	多态位点 Polymorphic loci	多态百分数/% Polymorphism percentage
E32-M48	1.9963	1.2604	0.1879	0.3211	269	99.63
E35-M48	1.9774	1.4286	0.2698	0.4234	260	97.74
E35-M60	1.9937	1.3928	0.2479	0.3944	158	99.37
E37-M49	1.9953	1.5302	0.3144	0.4765	213	99.53
E37-M59	2.0000	1.5961	0.3452	0.5156	246	100.00
E37-M60	1.9817	1.5571	0.3252	0.4890	214	98.17
E38-M48	1.9942	1.5507	0.3236	0.4881	172	99.42
E38-M49	1.9913	1.4182	0.2662	0.4199	229	99.13
E38-M60	1.9850	1.4554	0.2872	0.4473	263	98.50
E75-M55	1.9895	1.4117	0.2613	0.4130	283	98.95