赣南野生山金柑全基因组InDel标记开发及应用

doi:10.16420/j.issn.0513-353x.2024-0357

摘要/Abstract

摘要：

为探究赣南野生山金柑InDel特征，选取99份赣南野生山金柑进行全基因组重测序，进而筛选InDel变异位点以开发InDel标记。获得590.58 Mb有效片段数据，共检测到211 470个InDel，包括98 043个插入和113 427个缺失。从8 760条插入/缺少碱基数大于10的InDel标记中选择出180条在染色体上均匀分布的多态性标记，并开发了58对扩增条带良好的InDel引物。InDel标记聚类与全基因组重测序聚类结果一致，99份赣南野生山金柑分为既存在关联性又具有特异性的4个分支；根据每份种质含有的核心InDel标记位点数和等位基因数，确定了29份赣南野生山金柑为核心种质，基因组覆盖度达99.7%，其中夹湖、余坑和蔡坊是种质核心区域；结合全基因组重测序数据和核心InDel标记构建了种质的DNA指纹图谱，图谱信息包含植株编号、基因型、变异位点和农艺性状数据。研究明确了赣南野生山金柑InDel特征，筛选的InDel标记能够应用在群体结构分析、核心种质筛选以及DNA指纹图谱构建等方面。

关键词: 山金柑, InDel标记, 遗传多样性, 核心种质, DNA指纹图谱

Abstract:

To clarify the InDel characteristics of Gannan（Southern Jiangxi Province）wild Fortunella hindsii，99 Gannan wild F. hindsii were selected for whole genome resequencing，and then InDel variant sites were screened to develop InDel markers. A total of 590.58 Mb valid fragment data was obtained，and 211 470 InDels were detected，including 98 043 insertions and 113 427 deletions. From 8 760 InDel markers with Insertion or Deletion base numbers greater than 10 180 polymorphic markers with uniform distribution on chromosomes were selected，and 58 pairs of InDel primers with good amplification bands were developed. The findings of 58 pairs of InDel markers demonstrated a similar grouping pattern with the clustering results obtained from whole genome resequencing. Additionally，the 99 Gannan wild F. count of core InDel marker sites and alleles present in each germplasm，a total of 29 Gannan wild F. hindsii were selected as the core germplasm. These germplasm samples had a genome coverage of 99.7%. Among them，Jiahu，Yukeng，and Caifang were recognised as the core germplasm regions. The DNA fingerprint was created by integrating the entire genome resequencing data with the core InDel marker sites. This fingerprint contains coding information such as plant number，genotype，mutation locus，and agronomic trait data. This study aimed to elucidate the InDel characteristics of Gannan wild F. hindsii. The identified InDel markers have potential applications in population structure analysis，core germplasm screening，and DNA fingerprint construction.

Key words: Fortunella hindsii, InDel marker, genetic diversity, core germplasm, DNA fingerprinting

罗嗣芳, 严翔, 谢丽芳, 孙静贤, 郭紫晶, 张祖铭, 陈兆星. 赣南野生山金柑全基因组InDel标记开发及应用[J]. 园艺学报, 2025, 52(2): 267-278.

LUO Sifang, YAN Xiang, XIE Lifang, SUN Jingxian, GUO Zijing, ZHANG Zuming, CHEN Zhaoxing. Development and Application of Insertion-Deletion（InDel）Markers in Gannan wild Fortunella hindsii Based on Whole Genome Re-sequencing Data[J]. Acta Horticulturae Sinica, 2025, 52(2): 267-278.

图/表 9

参考文献 21

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序号 No.	采集地 Collection place	编号 ID	生境 Habitat	海拔/m Altitude
1 ~ 13	蔡坊 Caifang（CF）	CF001 ~ CF013	山谷 Valley	271.5
14 ~ 16	三百山 Sanbaishan（SBS）	SBS001 ~ SBS003	原始森林 Primary forest	417.8
17 ~ 20	张牯段zhangguduan（ZGD）	ZGD001 ~ ZGD004	人工移植，丘陵 Artificial plant，hills	315.4
21 ~ 23	里仁 Liren（LR）	LR001 ~ LR003	人工移植，盆景 Artificial plant，Bonsai	353.7
24 ~ 27	余坑Ⅰ Yukeng Ⅰ（YK Ⅰ）	YK Ⅰ 001 ~ YK Ⅰ 004	人工移植，山谷 Artificial plant，valley	277.8
28 ~ 36	余坑Ⅱ Yukeng Ⅱ（YK Ⅱ）	YK Ⅱ 001 ~ YK Ⅱ 009	人工移植，山谷 Artificial plant，valley	277.8
37 ~ 46	余坑Ⅲ Yukeng Ⅲ（YK Ⅲ）	YK Ⅲ 001 ~ YK Ⅲ 010	人工移植，山谷 Artificial plant，valley	277.8
47 ~ 48	余坑Ⅳ Yukeng Ⅳ（YK Ⅳ）	YK Ⅳ 001 ~ YK Ⅳ 002	人工移植，山谷 Artificial plant，valley	277.8
49 ~ 88	夹湖 Jiahu（JH）	JH001 ~ JH040	人工移植，丘陵 Artificial plant，hills	271.6
89 ~ 93	沙石 Shashi（SS）	SS001 ~ SS005	人工移植，山谷 Artificial plant，valley	149.8
94 ~ 96	崇义 Chongyi（CY）	CY001 ~ CY003	原始森林 Primary forest	360.1
97 ~ 99	于都 Yudu（YD）	YD001 ~ YD003	山林 Forest	343.2

序号 No.	采集地 Collection place	编号 ID	生境 Habitat	海拔/m Altitude
1 ~ 13	蔡坊 Caifang（CF）	CF001 ~ CF013	山谷 Valley	271.5
14 ~ 16	三百山 Sanbaishan（SBS）	SBS001 ~ SBS003	原始森林 Primary forest	417.8
17 ~ 20	张牯段zhangguduan（ZGD）	ZGD001 ~ ZGD004	人工移植，丘陵 Artificial plant，hills	315.4
21 ~ 23	里仁 Liren（LR）	LR001 ~ LR003	人工移植，盆景 Artificial plant，Bonsai	353.7
24 ~ 27	余坑Ⅰ Yukeng Ⅰ（YK Ⅰ）	YK Ⅰ 001 ~ YK Ⅰ 004	人工移植，山谷 Artificial plant，valley	277.8
28 ~ 36	余坑Ⅱ Yukeng Ⅱ（YK Ⅱ）	YK Ⅱ 001 ~ YK Ⅱ 009	人工移植，山谷 Artificial plant，valley	277.8
37 ~ 46	余坑Ⅲ Yukeng Ⅲ（YK Ⅲ）	YK Ⅲ 001 ~ YK Ⅲ 010	人工移植，山谷 Artificial plant，valley	277.8
47 ~ 48	余坑Ⅳ Yukeng Ⅳ（YK Ⅳ）	YK Ⅳ 001 ~ YK Ⅳ 002	人工移植，山谷 Artificial plant，valley	277.8
49 ~ 88	夹湖 Jiahu（JH）	JH001 ~ JH040	人工移植，丘陵 Artificial plant，hills	271.6
89 ~ 93	沙石 Shashi（SS）	SS001 ~ SS005	人工移植，山谷 Artificial plant，valley	149.8
94 ~ 96	崇义 Chongyi（CY）	CY001 ~ CY003	原始森林 Primary forest	360.1
97 ~ 99	于都 Yudu（YD）	YD001 ~ YD003	山林 Forest	343.2

染色体 Chromosome	插入 Insertion	缺失 Deletion	总计 Total	占比/% Proportion
Chr1	12 831	14 127	26 958	12.75
Chr2	13 704	14 709	28 413	13.44
Chr3	13 086	14 487	27 573	13.04
Chr4	6 321	11 316	17 637	8.34
Chr5	11 916	12 480	24 396	11.54
Chr6	8 223	11 244	19 467	9.20
Chr7	14 157	15 816	29 973	14.17
Chr8	8 598	9 360	17 958	8.49
Chr9	9 153	9 810	18 963	8.97
染色体骨架 Scaffold	54	78	132	0.06
总计Total	98 043	113 427	211 470	100.00

染色体 Chromosome	插入 Insertion	缺失 Deletion	总计 Total	占比/% Proportion
Chr1	12 831	14 127	26 958	12.75
Chr2	13 704	14 709	28 413	13.44
Chr3	13 086	14 487	27 573	13.04
Chr4	6 321	11 316	17 637	8.34
Chr5	11 916	12 480	24 396	11.54
Chr6	8 223	11 244	19 467	9.20
Chr7	14 157	15 816	29 973	14.17
Chr8	8 598	9 360	17 958	8.49
Chr9	9 153	9 810	18 963	8.97
染色体骨架 Scaffold	54	78	132	0.06
总计Total	98 043	113 427	211 470	100.00

染色体 Chromosome	引物名称 Primer name	物理位置 Location	正向引物序列（5′-3′） Forward primer sequences	反向引物序列（5′-3′） Reverse primer sequences
Chr.1	Fh1-01	345 790	ATGCGCCCATATCAACAAAC	CTCGACCTCACAAGGGTCAT
	Fh1-03	1 028 626	TGCGAACTCTCAAAGAGCAA	CGCATGAAAAGTACCGGTGT
	Fh1-05	2 758 295	TTTAGCTCACGTGCAAGTCC	GCGACAAAATTCAACAATGC
	Fh1-08	6 542 069	ATCAATGCCGAAAGAAATCG	GCGGGAAAACCAAATGTTCT
	Fh1-14	12 876 172	TTACCTCTGTGGGCATGCTT	CGGCAATGGAACTGATTTCT
	Fh1-18	20 696 997	ATGTTCTTGGGCATCCTCAC	CAGCAAATGCAACCATCAAG
	Fh1-20	28 766 849	AAATTGTGATTGCTGGAGGC	GGAGGACAGCTTCCGTATTG
Chr.2	Fh2-01	109 117	GCCAAGCTCGACTTCACCTA	TCTCTTGTCGTGTTGGGTTG
	Fh2-03	5 553 966	TTCAACACACCAATTTTAAGAGATTT	TTCTGCTTCGTCCCTTCTTC
	Fh2-05	8 613 056	GCATTTCCCACCAATGTTTT	TTTGGAGCAGGATTTGATTTG
	Fh2-08	15 929 836	TGATGGAGTTGAAGCAGCAC	ACAACAAAAACAAGGCCAGC
	Fh2-11	21 318 214	CATGGGACTAAAATTGGGACC	CCAGGGTTCTTCTAGACGGT
	Fh2-16	28 640 384	AAATCCAACGGCGTACTTGT	TCAACGGATAACTTCTCGACG
	Fh2-18	30 563 177	GCATGCCTTTTGGTTTCATT	ATGCTCAAATCGCCTTCATT
Chr.3	Fh3-01	1 382 984	TTTGGGATATGAAATAATCTTCTATGA	TCTTTCTTTTCCCCTTTTTGG
	Fh3-04	4 927 541	ACGAGCAGAAGAGCTGAAGG	GCATGGCAAAGTAAGCGAAT
	Fh3-07	8 409 496	ATCGCTTAAATTGCTAGGGC	GCTCCAGCAATCCTTCTTTG
	Fh3-11	13 680 425	CAGGTTGCAATGTTGGATTC	GGGTGGACCCAAGAATTGAT
	Fh3-15	16 819 522	CAGCAGATGTCAGGAAATGG	TTGAGTCCAAACTCCAACCTG
	Fh3-17	21 775 047	TTCTGCCGTTTTTGTGTGAG	ATACGGCTATCGGCACAATC
	Fh3-19	26 439 136	AAGCTTTCACCAAATGCACC	TTCCCTGTAGAAGCATTGGC
Chr.4	Fh4-04	4 839 437	ACAAAACAAGAGAATGGCCG	CACGAAAAAGCAGAATGCAA
	Fh4-12	12 857 077	CTACCCACACACGAGAAGCA	AAATCGGAAAGGCCAAAAGT
	Fh4-15	14 192 085	CGTTGCCGGAAAGTTTTCTA	TTTGCAAAAGTTTTTGGGCT
	Fh4-19	19 886 665	TCATCCGAACACGGTATTGA	GATCGTGGTAAAGAGGGCAA
Chr.5	Fh5-03	3 603 056	CCTTGGGCTTTGTTGTTTGT	CCCGAGGCTCTTACAGGAAT
	Fh5-07	6 518 497	ATTTGTTGCTTGATCCGAGG	GGATTGGCTCTCCACTTTGT
	Fh5-11	11 510 323	CGGGTGGGTTTGGTATTAAC	TAAGCACCCATCCAACACAA
	Fh5-14	19 373 991	AATTTGTTTGCCTGTACCCG	ATTCAGGCAGAGCTGGTTGT
	Fh5-18	25 493 920	AATCCATTAAACGATCGGCA	GTTTGTGGCAAATGAATCCC
	Fh5-20	28 017 235	TCACCGCAAAAACTCAACAA	ATGGCTCCCTCCTTTCATCT
Chr.6	Fh6-01	9 495	CCTGAATACCCCCGTCCTAT	CTCAGAACTGCTCCCCTCAC
	Fh6-06	6 836 044	GGAAATGTCTGAGCATGCAA	TTTTCCACCCACAACACAAA
	Fh6-10	10 384 324	TTTCATCCTCATCAAGTCCTCA	AGTGAAAACCGGACCACAAG
	Fh6-14	14 869 148	TAGTTTGAATTGCGAACCCC	TACACGCATGAGCTGAGAGC
	Fh6-17	18 459 550	AATTTTACCGGGCTCTCCTC	AACAACATAACTCGCTGCCA
	Fh6-19	21 169 068	CTGTTTTTCGGCGTTCAAA	CCTCCGATTGTGGTTCTGTT
Chr.7	Fh7-01	97 643	CCTTTGTGAGAGGGTAAGCG	GGCGCAATCATTTCTTTCAT
	Fh7-05	4 570 335	GGGCCAAAGAGTATTGATTTTT	CCACCACCAACTGACCTTTA
	Fh7-07	8 179 352	GATTCCTTCTCTCGTGTGGC	TGGAACTTGGAACAACACCA
	Fh7-09	11 357 338	AAACCGAAACCAAACCCAAT	TCTCTAATGAGGGTCGGGGT
	Fh7-10	14 093 476	ATTCAGGTCGATATACGGCG	ATCGTCCAAAATGGAGCAAC
	Fh7-11	19 572 019	CAGTACCACAGCAAGACGGA	ACTTGGTTAATCCCCGAACC
	Fh7-14	25 270 436	GGGCCAAGCTAGCACACTTA	CTTATTGTCGTAGGCCGCTC
	Fh7-16	28 329 053	GGCAATAGCCGTTCCATCTA	AATTTTCAGTTTGAGGCCGA
	Fh7-19	31 726 654	CGCCAACTCAGAGACATTGA	ATCGCTTGTGAAGCTTTGCT
	Fh7-20	32 198 988	AGGCCCATGTAGAAAGCTGA	TTGCACGTGCTTTTCCATAG
Chr.8	Fh8-03	537 740	TTGAGCAAAGATGGCGAATA	TGCCAAGAGCAACAGGAATA
	Fh8-06	5 852 004	GCAAGCATGTGCATTTAGGA	AATGGCTTCATGTCAAAGGG
	Fh8-09	8 990 917	GGAGGTTGCAAATATGGCTG	ACAAGGTGGCTGACCGTTAC
	Fh8-16	14 784 483	AATCACGCCAAGAAATCCAC	TCAACCGCACCTATAACCAA
	Fh8-19	19 111 826	TCAAACGTCCACGTTGTTCT	GGAGAAGGTCTCCTCATCCC
	Fh8-20	20 640 431	GTTGTGTTGCTCTGGGGTTT	CCACTGTGTATCTGGGTGGA
Chr.9	Fh9-01	2 559 855	TTTTGAACCATTGATTTCAAGC	TGCCCTTTTAAATCTCCAGTT
	Fh9-09	9 980 616	CATGCAGGCATACACAGTGA	GATGGGGCGAAAGAGTACAA
	Fh9-11	10 172 322	TACATAGGCATTTCCCCTGC	TTGGGAGATGCTTTCAGAGG
	Fh9-15	16 276 097	ATATGAGTTTCGGTGAGCCG	TCTGGATAGCCAACCAAGTAAAA
	Fh9-19	18 330 523	GACTCTCCCACTGGACCAAA	GTGCACATTATTCATTGGCG