西藏石榴野生群体的SSR遗传多样性分析

doi:10.12302/j.issn.1000-2006.202105011

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南京林业大学学报（自然科学版） ›› 2022, Vol. 46 ›› Issue (3): 127-133.doi: 10.12302/j.issn.1000-2006.202105011

西藏石榴野生群体的SSR遗传多样性分析

葛大朋¹(), 任媛¹, 赵俊², 王玉婷², 刘学庆³, 苑兆和¹^,^*()

1.南京林业大学,南方现代林业协同创新中心,南京林业大学林学院,江苏南京 210037
2.西藏自治区林木科学研究院,西藏拉萨 850000
3.山东省烟台市农业科学研究院,山东烟台 265500

收稿日期:2021-05-10 接受日期:2021-07-24 出版日期:2022-05-30 发布日期:2022-06-10
通讯作者: 苑兆和
基金资助:
西藏自治区自然科学基金项目(XZ2019 ZR G-153);国家自然科学基金项目(31901341);江苏省自然科学基金青年基金项目(BK20180768);南京林业大学高层次人才科研启动基金项目(GXL2014070);南京林业大学高层次人才科研启动基金项目(GXL2018032);江苏高校优势学科建设工程项目(PAPD)

Genetic diversity among wild populations of pomegranate in Tibet by SSR analyses

GE Dapeng¹(), REN Yuan¹, ZHAO Jun², WANG Yuting², LIU Xueqing³, YUAN Zhaohe¹^,^*()

1. Co-innovation Center for Sustainable Forestry in Southern China, College of Forestry, Nanjing Forestry University, Nanjing 210037, China
2. Institute of Forestry Science of Tibet Autonomous Region, Lhasa 850000, China
3. Yantai Academy of Agricultural Science, Yantai 265500, China

Received:2021-05-10 Accepted:2021-07-24 Online:2022-05-30 Published:2022-06-10
Contact: YUAN Zhaohe

摘要/Abstract

摘要：

【目的】调查西藏地区野生石榴种质资源,分析西藏野生石榴群体的遗传多样性和遗传结构,为野生石榴资源的保护和利用提供理论依据。【方法】使用13对SSR引物对3个自然群体共42份西藏地区野生石榴种质资源材料DNA进行PCR扩增,毛细管电泳检测扩增片段长度,使用GenAlEx和Arlequin等软件对SSR数据进行分析。【结果】13对引物共检测到44个等位基因,平均3.385个,引物的平均有效等位基因数(N_e)、香农信息指数(I)、期望杂合度(H_e)和多态信息量(PIC)分别为1.971、0.771、0.481和0.393。3个野生群体的平均有效等位基因数(N_e)、平均香农信息指数(I)和平均期望杂合度(H_e)分别为1.867、0.646和0.421,林芝b(LZb)群体的遗传多样性水平高于其他2个群体。AMOVA分析表明,群体内遗传变异高达88.43%,3个群体间的遗传分化系数(F_st)为0.116。种质聚类分析将供试种质划分为3个亚群,结果与种质地理来源具有一定关联性。遗传结构分析显示西藏野生石榴有4个可能的基因库来源。【结论】13对SSR引物可用于西藏野生石榴种质的遗传多样性等研究。西藏野生石榴种质的遗传变异主要存在于群体内;林芝b(LZb)群体遗传多样性最高,遗传结构复杂,且含有最多的野生种质采样点,可予以优先保护。

关键词: 石榴野生群体, SSR, 遗传多样性, 遗传结构, 西藏

Abstract:

【Objective】Wild pomegranate (Punica granatum) accessions were found in Tibet. To facilitate the conservation and use of wild pomegranate germplasm, the genetic diversity and population structure of three wild pomegranate populations from Tibet were studied.【Method】Primers for 13 SSR loci were used for PCR amplification of 42 wild individuals from from populations, and the amplified fragment length was determined using capillary electrophoresis. SSR data were analyzed using GenAlEx and Arlequin softwares.【Result】A total of 44 alleles were detected using 13 pairs of primers with an average of 3.835 alleles per locus. The mean values for the effective number of alleles (N_e), Shannon’s Information Index (I), expected heterozygosity (H_e) and the polymorphism information content (PIC) per locus were 1.971, 0.771, 0.481 and 0.393, respectively. The mean N_e, I and H_e of three wild populations were 1.867, 0.646 and 0.421, respectively. In the three populations of wild pomegranate accessions, the genetic diversity of the LZb population was higher than that of the CD and LZa populations. The AMOVA results showed that the percentage of genetic variation within populations was 88.43%, whereas the variance among populations was 11.57%. The F_st among the three wild populations was 0.116. Cluster analysis grouped the 42 accessions into three distinct clusters. Cluster analysis was found to be correlated with geographic origin. The most appropriate number of subpopulations was found to be K=4 using structure analysis.【Conclusion】A total of 13 SSR primers were found to be suitable for genetic diversity analysis of wild pomegranate accessions in Tibet. The genetic variance of wild pomegranate accessions in Tibet was mainly within populations. With having the highest genetic diversity and rich sampling sites, the LZb population merits should be a conservation priority.

Key words: wild populations of pomegranate (Punica granatum), SSR, genetic diversity, genetic structure, Tibet

中图分类号:

S665.4
S718

葛大朋,任媛,赵俊,等. 西藏石榴野生群体的SSR遗传多样性分析[J]. 南京林业大学学报（自然科学版）, 2022, 46(3): 127-133.

GE Dapeng, REN Yuan, ZHAO Jun, WANG Yuting, LIU Xueqing, YUAN Zhaohe. Genetic diversity among wild populations of pomegranate in Tibet by SSR analyses[J].Journal of Nanjing Forestry University (Natural Science Edition), 2022, 46(3): 127-133.DOI: 10.12302/j.issn.1000-2006.202105011.

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群体 population	采样点 site	北纬/(°) latitude (N)	东经/(°) longitude (E)	海拔/m altitude
昌都 CD	昌都市八宿县林卡乡叶巴村	29.879 3	97.379 9	2 661
	昌都市左贡县东坝乡普卡村	29.852 7	97.422 3	2 673
	昌都市左贡县东坝乡军拥村	29.856 8	97.424 3	2 699
林芝a LZa	林芝市察隅县古拉乡	29.020 5	97.952 5	2 374
林芝a LZa	林芝市察隅县古拉乡安巴村	29.040 5	97.982 8	2 322
林芝b LZb	林芝市察隅县察瓦龙乡珠拉村	28.736 4	98.439 4	2 125
	林芝市察隅县察瓦龙乡格布村	28.611 2	98.451 9	2 299
	林芝市察隅县察瓦龙乡则拉村	28.492 0	98.454 5	2 053
	林芝市察隅县察瓦龙乡邓许村	28.443 3	98.457 1	1 798
	林芝市察隅县察瓦龙乡昌西村	28.371 7	98.452 5	1 791

引物名称 primer name	重复单元 repeat motif	正向引物(5'-3') forward primer	反向引物(5'-3') reverse primer	来源 source
48	(AAAG)11	AAACGGGTGACTAAGAAATTTT	TTCACCCTCACCTAGACGACAT	自主开发
59	(TACA)9	TGCAGCAGTTTCTCTTCT	GTGGTGTCTGACGTGCGA	自主开发
73	(TGTA)9	TCTTCCAAAAGGGCGAAT	GTGGAGTGCATCAACCAA	自主开发
86	(TATC)9	TTCCAGCAAGGCAAGTGTTT	ATACGCCGGTTTTTCGATCT	自主开发
99	(CGAA)6	CTAAGGGATTTTGTTGGC	GCACAAAGTCAAAACACA	自主开发
136	(TTTA)5	ACAAAAAGACGGGAACCT	AGACCTCATAAAACTGGG	自主开发
140	(GTC)6	ATAGGCAATAACAAAGGT	AACCCGCAATCGACTTTA	自主开发
149	(AGGA)5	TCCCCAGAACTGAACACA	GCCAATGATAATCCACAT	自主开发
dbz4	CT	CATCAGAATCGTCCCCTTGTG	CAGAGAGAAGAAGAGAGACCGAGC	文献[15]
M06	(CTT)26	TGGTTAGGCATGCAAGAGTG	GAGCTGGTGATTTCATTGGG	文献[6]
M07	(TTC)25	TAGCCCCATTTTGCTGATTC	GTCGCGTGAAATTCCCTAAA	文献[6]
tsh4	(TATC)9	AAAGCATGCGAAAGGATGAT	CAATGCCAATGTTACGGATG	文献[4]
tsh6	(GTCT)7	CGATGTGTACAGTTGTGGGC	CCGTTTTAGCTCCAGTCTGC	文献[4]

位点 locus	N_a	N_e	I	H_o	H_e	F	多态信息含量 PIC
48	3	1.999	0.737	0.429	0.500	0.142	0.386
59	2	1.600	0.562	0.167	0.375	0.556	0.305
73	3	2.119	0.817	0.429	0.528	0.188	0.421
86	4	1.504	0.621	0.143	0.335	0.574	0.299
99	3	1.925	0.718	0.548	0.480	-0.140	0.376
136	2	1.774	0.628	0.643	0.436	-0.474	0.341
140	2	1.930	0.675	0.381	0.482	0.209	0.366
149	2	1.747	0.619	0.429	0.427	-0.003	0.336
dbz4	2	1.866	0.657	0.488	0.464	-0.051	0.356
M06	6	2.197	0.916	0.405	0.545	0.257	0.442
M07	6	2.040	1.036	0.214	0.510	0.580	0.469
tsh4	3	2.108	0.815	0.561	0.526	-0.067	0.421
tsh6	6	2.813	1.227	0.500	0.645	0.224	0.591
均值mean	3.385	1.971	0.771	0.410	0.481	0.154	0.393

群体 population	采样点数量 sampling site	种质数量 accession	N_a	N_e	I	H_o	H_e	F
CD	3	20	2.308	1.817	0.600	0.493	0.395	-0.217
LZa	2	5	1.769	1.695	0.513	0.077	0.365	0.774
LZb	5	17	3.077	2.091	0.824	0.412	0.504	0.176
均值 mean	3.3	14	2.385	1.867	0.646	0.327	0.421	0.212

变异来源 source	自由度 df	离差平方和 SS	变异方差分量 variance components	方差分量占比/% percentage of variation
群体间 among populations	2	24.868	0.381	11.57
群体内 within populations	81	235.799	2.911	88.43
总计 total	83	260.667	3.292	100.00
F_st	0.116

西藏石榴野生群体的SSR遗传多样性分析

Genetic diversity among wild populations of pomegranate in Tibet by SSR analyses

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群体1 population 1	群体2 population 2	F_st	N_m		种质数accessions
群体1 population 1	群体2 population 2	F_st	N_m		群体1 population 1		群体2 population 2
CD	LZa	0.229		0.840		20		5
CD	LZb	0.086		2.662		20		17
LZa	LZb	0.060		3.924		5		17

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群体 population	红色类群 Q1	淡绿色类群 Q2	黄色类群 Q3	天蓝色类群 Q4	混合类群 mix	总和 total
CD	0	5	0	12	3	20
LZa	0	0	3	0	2	5
LZb	8	0	0	4	5	17
总计 total	8	5	3	16	10	42