Genetic Diversity Analysis about Astringency of Walnut by SSR Molecular Marker

doi:10.7668/hbnxb.20193708

Abstract

Abstract:

To study the genetic diversity of astringency in walnut germplasm resources,118 germplasm resources from 4 different astringency walnut populations were used as materials,which was studied by using SSR capillary electrophoresis fluorescence labeling technology,and the dendrogram of 4 populations was constructed.The results showed that a total of 93 allelic variants were detected by 12 pairs of primers,ranging from 2 to 18,with an average of 7.75 alleles detected by each pair of SSR primers.Polymorphism information content (PIC) ranged from 0.357 9 to 0.785 2,with an average of 0.541 1.The number of polymorphic sites (Np) was 91,the percentage of polymorphic sites (PPB) was 97.85%,the number of observed alleles (Na) was 1.978 5,the effective number of alleles (Ne) was 1.198 5,Shannon's information index (I) was 0.209 7,Nei's diversity index (H) was 0.126 3,number of total genetic diversity index (Ht) was 0.126 7 and intra-population genetic diversity index (Hs) was 0.122 2,indicating that the genetic diversity and variation of the four walnut populations were not high.The coefficient of genetic differentiation among populations was 0.035 5,indicating that the genetic variation within populations was 96.45%.The percentage of polymorphic loci in the slightly astringent population was the largest,which was 72.04%,indicating that the slightly astringent population had the richest genetic diversity among the four populations.UPGMA cluster analysis showed that the genetic identity between four populations ranged from 0.989 8 to 0.997 1,indicating that the genetic distance among four populations was close.The four populations can be divided into two groups at the coefficient of 0.993 4.The first group included unastringent,slightly astringent and more astringent populations,with the astringent populations as a separate group.By calculating the gene frequency of 93 locis in different populations,41 locis of 12 pairs of SSR primers were found to be able to distinguish the resources with different levels of astringency.SSR capillary electrophoresis fluorescence labeling technology was used to analyze the genetic diversity related to astringency in 4 walnut populations.According to the transcriptomic sequencing results,12 pairs of primers were selected for SSR analysis.A total of 93 SSR locis were amplified,and the percentage of polymorphic sites was 97.85%,indicating that the primers had high polymorphism and were suitable for genetic diversity analysis of walnut,which can be used for subsequent studies on genetic diversity and breeding of astringency in walnut.

Key words: Walnut, SSR, Astringency, Genetic diversity, Capillary electrophoresis

LIU Jing, LI Yang, LIU Jinli, ZHANG Xinfang, BAI Zhongkui, YU Qiuxiang. Genetic Diversity Analysis about Astringency of Walnut by SSR Molecular Marker[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(S1): 23-30. doi: 10.7668/hbnxb.20193708.

/ / Recommend / Download Citations

URL: http://www.hbnxb.net/EN/10.7668/hbnxb.20193708

http://www.hbnxb.net/EN/Y2024/V39/IS1/23

Figures/Tables 7

References 32

[1]	刘警, 于秋香, 李扬, 李义红, 张家齐, 白仲奎. 我国核桃生产的现状、问题及发展对策[J]. 北方果树, 2020(6):38—41.doi:10.16376/j.cnki.bfgs.2020.06.016.
	Liu J, Yu Q X, Li Y, Li Y H, Zhang J Q, Bai Z K. Present situation,problems and development countermeasures of walnut production in China[J]. Northern Fruits, 2020(6):38—41.
[2]	傅本重, 邹路路, 朱洁倩, 魏蜜, 杨新河, 王立华, 李国元. 中国核桃生产现状与发展思路[J]. 江苏农业科学, 2018, 46(18):5—8.doi:10.15889/j.issn.1002-1302.2018.18.002.
	Fu B Z, Zou L L, Zhu J Q, Wei M, Yang X H, Wang L H, Li G Y. Production status and development of Chinese walnut[J]. Jiangsu Agricultural Sciences, 2018, 46(18):5—8.
[3]	裴东, 鲁新政. 中国核桃种质资源[M]. 北京: 中国林业出版社, 2011.
	Pei D, Lu X Z. Walnut germplasm resources in China[M]. Beijing: China Forestry Publishing House, 2011.
[4]	肖良俊, 陈少瑜, 宁德鲁, 吴涛, 贺娜. 滇西北深纹核桃(Juglans sigillata)种质资源遗传多样性研究[J]. 云南农业大学学报(自然科学), 2018, 33(4):597—604.doi:10.12101/j.issn.1004-390X(n).201804049.
	Xiao L J, Chen S Y, Ning D L, Wu T, He N. Study on the genetic diversity of germplasm resources of Juglans sigillata in northwest Yunnan[J]. Journal of Yunnan Agricultural University(Natural Science), 2018, 33(4):597—604.
[5]	刘晓丽, 何天明, 张美勇, 张立杰, 慈志娟, 张春雨, 陈学森. 核桃SSR反应体系的优化[J]. 果树学报, 2007, 24(2):140—145.doi:10.3969/j.issn.1009-9980.2007.02.003.
	Liu X L, He T M, Zhang M Y, Zhang L J, Ci Z J, Zhang C Y, Chen X S. Optimization of SSR analysis system in walnut[J]. Journal of Fruit Science, 2007, 24(2):140—145.
[6]	包文泉, 乌云塔娜, 杜红岩, 李铁柱, 刘慧敏, 王淋, 白玉娥. 基于SSR标记的西藏光核桃群体遗传多样性和遗传结构分析[J]. 林业科学, 2018, 54(2):30—41.doi:10.11707/j.1001-7488.20180204.
	Bao W Q, Wu Y, Du H Y, Li T Z, Liu H M, Wang L, Bai Y E. Genetic diversity and population structure of amygdalus mira in the Tibet Plateau in China based on SSR markers[J]. Scientia Silvae Sinicae, 2018, 54(2):30—41.
[7]	周于波, 朱鹏, 龚伟, 王景燕, 闫思宇, 吴开志, 吴春艳, 李海平, 龚毅红, 段琼. 基于SSR标记的川西南泡核桃良种DNA指纹图谱构建及聚类分析[J]. 分子植物育种, 2018, 16(17):5683—5689.doi:10.13271/j.mpb.016.005683.
	Zhou Y B, Zhu P, Gong W, Wang J Y, Yan S Y, Wu K Z, Wu C Y, Li H P, Gong Y H, Duan Q. DNA fingerprinting establishment and clustering analysis of Juglans sigllata improved variety from southwestern Sichuan based on SSR markers[J]. Molecular Plant Breeding, 2018, 16(17):5683—5689.
[8]	刘雨霞, 张玲, 张小军, 杨笑, 杨春, 曲进, 刘群龙. 基于电子舌技术分类评价核桃内种皮的口感品质[J]. 食品与发酵工业, 2020, 46(19):258—263.doi:10.13995/j.cnki.11-1802/ts.024127.
	Liu Y X, Zhang L, Zhang X J, Yang X, Yang C, Qu J, Liu Q L. Classification and evaluation of walnut kernel pellicle taste quality based on electronic tongue technology[J]. Food and Fermentation Industries, 2020, 46(19):258—263. doi: 10.13995/j.cnki.11-1802/ts.024127
[9]	罗晓文, 刘敏, 齐晓花, 徐强, 陈学好. 果实涩味分子研究进展[J]. 分子植物育种, 2013, 11(5):647—656.doi:10.3969/mpb.011.000647.
	Luo X W, Liu M, Qi X H, Xu Q, Chen X H. Molecular research progress in fruit astringent[J]. Molecular Plant Breeding, 2013, 11(5):647—656.
[10]	Shahidi F, Naczk M. Phenolics in food and nutraceuticals[J]. Phenolics in Food & Nutraceuticals, 2004, 13(3):12—15.
[11]	张泽生, 王霄然, 王田心, 秦程广, 徐梦莹, 王春龙, 郭擎. 核桃内种皮提取物的体内抗氧化活性研究[J]. 中国食品添加剂, 2017(1):110—114.doi:10.3969/j.issn.1006-2513.2017.01.010.
	Zhang Z S, Wang X R, Wang T X, Qin C G, Xu M Y, Wang C L, Guo Q. Antioxidant activity of phenolic extracts from walnut (Juglans regia L.) kernel pellicle in vivo[J]. China Food Additives, 2017(1):110—114.
[12]	Shimoda H, Tanaka J, Kikuchi M, Fukuda T, Ito H, Hatano T, Yoshida T. Walnut polyphenols prevent liver damage induced by carbon tetrachloride and d-galactosamine:hepatoprotective hydrolyzable tannins in the kernel pellicles of walnut[J]. Journal of Agricultural and Food Chemistry, 2008, 56(12):4444—4449.doi:10.1021/jf8002174.
[13]	Carvalho M, Ferreira P J, Mendes V S, Silva R, Pereira J A, Jerónimo C, Silva B M. Human cancer cell antiproliferative and antioxidant activities of Juglans regia L.[J]. Food and Chemical Toxicology, 2010, 48(1):441—447.doi:10.1016/j.fct.2009.10.043. pmid: 19883717
[14]	Anderson K J, Teuber S S, Gobeille A, Cremin P, Waterhouse A L, Steinberg F M. Walnut polyphenolics inhibit in vitro human plasma and LDL oxidation[J]. The Journal of Nutrition, 2001, 131(11):2837—2842.doi:10.1093/jn/131.11.2837.
[15]	Choi J G, Park G, Kim H G, Oh D S, Kim H, Oh M S. In vitro and in vivo neuroprotective effects of walnut (Juglandis semen) in models of Parkinson's disease[J]. International Journal of Molecular Sciences, 2016, 17(1):108.doi:10.3390/ijms17010108.
[16]	Pereira J A, Oliveira I, Sousa A, Ferreira I C F R, Bento A, Estevinho L. Bioactive properties and chemical composition of six walnut (Juglans regia L.) cultivars[J]. Food and Chemical Toxicology, 2008, 46(6):2103—2111.doi:10.1016/j.fct.2008.02.002.
[17]	朱亚新, 陈朝银, 赵声兰. 核桃多酚对酪氨酸酶的作用[J]. 食品工业科技, 2014, 35(22):382—385,391.doi:10.13386/j.issn1002-0306.2014.22.075.
	Zhu Y X, Chen C Y, Zhao S L. Effect of polyphenols from walnut on tyrosinase[J]. Science and Technology of Food Industry, 2014, 35(22):382—385,391.
[18]	Qamar W, Sultana S. Polyphenols from Juglans regia L.(walnut) kernel modulate cigarette smoke extract induced acute inflammation,oxidative stress and lung injury in Wistar rats[J]. Human & Experimental Toxicology, 2011, 30(6):499—506.doi:10.1177/0960327110374204.
[19]	Fukuda T, Ito H, Yoshida T. Effect of the walnut polyphenol fraction on oxidative stress in type 2 diabetes mice[J]. BioFactors, 2004, 21(1/2/3/4):251—253.doi:10.1002/biof.552210148.
[20]	Nishiyama S, Onoue N, Kono A, Sato A, Ushijima K, Yamane H, Tao R, Yonemori K. Comparative mapping of the ASTRINGENCY locus controlling fruit astringency in hexaploid persimmon (Diospyros kaki Thunb.) with the diploid D.lotus reference genome[J]. The Horticulture Journal, 2018, 87(3):315—323.doi:10.2503/hortj.okd-140..
[21]	Jamil W, Wu W, Ahmad M, Zhu Q G, Liu X F, Jin R, Yin X R. High-CO₂/hypoxia-modulated NAC transcription factors involved in de-astringency of persimmon fruit[J]. Scientia Horticulturae, 2019, 252:201—207.doi:10.1016/j.scienta.2019.03.018.
[22]	陈佳歆, 周沫, 毕金峰, 李旋, 郭崇婷, 陈芹芹, 辛广. CO₂脱涩对柿果理化特性、酚类成分及抗氧化能力的影响[J]. 食品科学, 2019, 40(13):28—35.doi:10.7506/spkx1002-6630-20181023-265.
	Chen J X, Zhou M, Bi J F, Li X, Guo C T, Chen Q Q, Xin G. Impact of CO₂ de-astringency on physicochemical properties,phenolic compounds and antioxidant capacities of persimmon fruit[J]. Food Science, 2019, 40(13):28—35.
[23]	俞文君, 金强, 李根, 张锐, 吴翠云, 王新建, 于军. 基于果实苦涩味新疆核桃资源遗传多样性分析[J]. 食品工业科技, 2020, 41(13):234—240.doi:10.13386/j.issn1002-0306.2020.13.037.
	Yu W J, Jin Q, Li G, Zhang R, Wu C Y, Wang X J, Yu J. Genetic diversity analysis of walnut germplasm resources based on bitter and astringent of fruits in Xinjiang[J]. Science and Technology of Food Industry, 2020, 41(13):234—240.
[24]	陈锦永, 靳路真, 程大伟, 顾红, 张威远, 张洋, 郭西智, 方金豹. 水果涩味研究进展[J]. 果树学报, 2016, 33(12):1556—1566.doi:10.13925/j.cnki.gsxb.20160275.
	Chen J Y, Jin L Z, Cheng D W, Gu H, Zhang W Y, Zhang Y, Guo X Z, Fang J B. Research progress on fruit astringency[J]. Journal of Fruit Science, 2016, 33(12):1556—1566.
[25]	李露双, 董文慧, 丁兴萃, 章志远, 孙春娃, 蔡函江. 麻竹笋转录组测序及苦涩味物质合成基因差异表达分析[J]. 林业科学研究, 2018, 31(4):38—46.doi:10.13275/j.cnki.lykxyj.2018.04.006.
	Li L S, Dong W H, Ding X C, Zhang Z Y, Sun C W, Cai H J. Transcriptome sequencing and differential expression analysis of bitter and astringent substances biosynthesis related gene in Dendrocalamus latiflorus[J]. Forest Research, 2018, 31(4):38—46.
[26]	Colaricˇ M, Štampar F, Hudina M, Solar A. Sensory evaluation of different walnut cultivars (Juglans regia L.)[J]. Acta Agriculturae Slovenica, 2006, 87(2):403—413.doi:10.14720/aas.2006.87.2.15118.
[27]	谢倩, 张诗艳, 叶清华, 王威, 陈清西. 鲜食橄榄发育成熟过程中多酚及相关酶活性的动态变化[J]. 果树学报, 2019, 36(6):774—784.doi:10.13925/j.cnki.gsxb.20180524.
	Xie Q, Zhang S Y, Ye Q H, Wang W, Chen Q X. Dynamic changes of polyphenols and related enzymes activity during the development and maturation of Chinese olive(Canarium album L.)[J]. Journal of Fruit Science, 2019, 36(6):774—784.
[28]	黄勇. 基于SRAP分子标记的小果油茶遗传多样性分析[J]. 林业科学, 2013, 49(3):43—50.doi:10.11707/j.1001-7488.20130306.
	Huang Y. Analysis of camellia meiocarpa genetic diversity based on SRAP markers[J]. Scientia Silvae Sinicae, 2013, 49(3):43—50.
[29]	Wang H, Pan G, Ma Q G, Zhang J P, Pei D. The genetic diversity and introgression of Juglans regia and Juglans sigillata in Tibet as revealed by SSR markers[J]. Tree Genetics & Genomes, 2014, 11(1):1.doi:10.1007/s11295-014-0804-3.
[30]	高玉娜, 赵登超, 孙永泉, 刘清华, 侯立群. 核桃属材用核桃SSR标记的遗传多样性分析[J]. 西北农业学报, 2011, 20(9):129—134.doi:10.3969/j.issn.1004-1389.2011.09.026.
	Gao Y N, Zhao D C, Sun Y Q, Liu Q H, Hou L Q. SSR analysis of genetic diversity of walnut used for timber[J]. Acta Agriculturae Boreali-occidentalis Sinica, 2011, 20(9):129—134.
[31]	刘朝斌, 张文辉. 核桃种质资源遗传多样性的SSR分析[J]. 西北植物学报, 2012, 32(6):1118—1125.doi:10.3969/j.issn.1000-4025.2012.06.008.
	Liu C B, Zhang W H. SSR analysis of genetic diversity among walnuts germplasm[J]. Acta Botanica Boreali-Occidentalia Sinica, 2012, 32(6):1118—1125.
[32]	郭计华, 程敏, 苗贵东, 夏可灿, 张培, 张万芹, 蔺维成, 王颖娟. 基于ISSR标记的贵州山核桃遗传多样性分析[J]. 种子, 2022, 41(1):84—88,封2.doi:10.16590/j.cnki.1001-4705.2022.01.084.
	Guo J H, Cheng M, Miao G D, Xia K C, Zhang P, Zhang W Q, Lin W C, Wang Y J. Genetic diversity analysis of Carya cathayensis in Guizhou based on ISSR markers[J]. Seed, 2022, 41(1):84—88,封2.

涩味等级 Astringency grade	品种来源 Origin	资源 Resource
不涩	新疆	2-21、2-27、4-3
Unastringent	河北	魁香、台营2、旮旯铺、3-17、5-39、D1、D4
	辽宁	礼品2号、辽宁10号、寒丰
	山东	花生1号、丰香、A19
	河南	中核2号
	山西	晋龙1号
	日本	43号心形
稍涩	新疆	新光、扎木台3号、新早丰、1-41、2-7、2-17、2-19、2-35、3-30、4-22、4-26、6-3、6-5、6-21、
Slightly astringent		6-29、6-33
	河北	绿岭、早硕、龙珠、硕香、昌黎2、昌黎3、昌黎4、昌黎5、台营3、台营4、SJL、Y1、1-32、1-38、2-15、
		2-16、2-40、3-10、3-12、3-22、3-26、3-34、3-39、5-28、5-30、5-35、5-36、5-37、5-42、6-1
	辽宁	辽宁5号、辽宁6号、辽宁7号、礼品1号、辽瑞丰
	山东	华山5号
	朝鲜	安边2号
	日本	清香
较涩	新疆	1-43、2-37、2-39、3-1、4-5、4-15、6-25、6-43
More astringent	河北	西岭、台营1、1-34、2-8、2-12、2-14、2-20、2-32、2-34、3-5、3-7、3-20、3-21、3-29、3-32、3-33、
		4-10、5-9、5-12、5-34
	辽宁	辽宁1号、辽宁4号、10901优系
	山东	B76、鲁果6号、岱香
	北京	美香
	河南	绿波
涩	新疆	2-25、2-29、4-29、6-7、纸皮3号
Astringent	河北	2-22、2-42
	山西	金薄丰
	河南	中核3号

涩味等级 Astringency grade	品种来源 Origin	资源 Resource
不涩	新疆	2-21、2-27、4-3
Unastringent	河北	魁香、台营2、旮旯铺、3-17、5-39、D1、D4
	辽宁	礼品2号、辽宁10号、寒丰
	山东	花生1号、丰香、A19
	河南	中核2号
	山西	晋龙1号
	日本	43号心形
稍涩	新疆	新光、扎木台3号、新早丰、1-41、2-7、2-17、2-19、2-35、3-30、4-22、4-26、6-3、6-5、6-21、
Slightly astringent		6-29、6-33
	河北	绿岭、早硕、龙珠、硕香、昌黎2、昌黎3、昌黎4、昌黎5、台营3、台营4、SJL、Y1、1-32、1-38、2-15、
		2-16、2-40、3-10、3-12、3-22、3-26、3-34、3-39、5-28、5-30、5-35、5-36、5-37、5-42、6-1
	辽宁	辽宁5号、辽宁6号、辽宁7号、礼品1号、辽瑞丰
	山东	华山5号
	朝鲜	安边2号
	日本	清香
较涩	新疆	1-43、2-37、2-39、3-1、4-5、4-15、6-25、6-43
More astringent	河北	西岭、台营1、1-34、2-8、2-12、2-14、2-20、2-32、2-34、3-5、3-7、3-20、3-21、3-29、3-32、3-33、
		4-10、5-9、5-12、5-34
	辽宁	辽宁1号、辽宁4号、10901优系
	山东	B76、鲁果6号、岱香
	北京	美香
	河南	绿波
涩	新疆	2-25、2-29、4-29、6-7、纸皮3号
Astringent	河北	2-22、2-42
	山西	金薄丰
	河南	中核3号

引物名称 Primers	正向引物(5'—3') Forward primers(5'—3')	反向引物(5'—3') Reverse primers(5'—3')	扩增物大小/bp The size of fragment	等位基因数 No. of alleles
PAL-1	F:CAGCAGCGACTGGGTCATAA	R:AGCATGGGACTTTTCCACCTATA	322~327	4
ANR-6	F:CCACCTAAGAAGCCACCGTT	R:GGCTCTCATACTTTTGCGATGC	101~141	11
UFGT-6	F:TTGGCTGGACGCGATAATGA	R:ATCTTCGACGGTTGGGCAAT	237~278	11
4CL	F:TCGAGACGTCCAGATCCAGT	R:CGAGGATGGGTCGCTTTCTT	131~156	4
DFR-2	F:GCACCGTTCGTTTATACCGC	R:GGTGGCCACCGAGAAATGTA	147~189	18
C4H	F:TCATTCGTGCCTATGTCGATACT	R:GCCTTTGTGCTCGACAGTTT	260~294	5
F3'H	F:GCCAAAGCCAAAGAAGAGGG	R:GCTTCAACAACCTTGCCATCA	385~465	8
UFGT-1	F:TCCCTCGATACATGTTGGCA	R:GCACATCTCACCACTCCTCC	288~299	8
CHS	F:GAGAAGTTCAAGCGCATGTGT	R:CCTTGGTGGCTGCTTCTTTG	484~518	6
ANR-3	F:TTGGTGGACGCTTGGTGATT	R:AGCCTCCACTCCGATCTCTT	222~258	11
ANR-5	F:CAGTAGAGTCTGCGTCACGG	R:ACTTTGGATGTGTCGCCTGT	282~311	5
FHT-4	F:CGAGTTGACACCCTCCACTC	R:ACGAAGCTTTTCCTCGGGAG	546~548	2

引物名称 Primers	正向引物(5'—3') Forward primers(5'—3')	反向引物(5'—3') Reverse primers(5'—3')	扩增物大小/bp The size of fragment	等位基因数 No. of alleles
PAL-1	F:CAGCAGCGACTGGGTCATAA	R:AGCATGGGACTTTTCCACCTATA	322~327	4
ANR-6	F:CCACCTAAGAAGCCACCGTT	R:GGCTCTCATACTTTTGCGATGC	101~141	11
UFGT-6	F:TTGGCTGGACGCGATAATGA	R:ATCTTCGACGGTTGGGCAAT	237~278	11
4CL	F:TCGAGACGTCCAGATCCAGT	R:CGAGGATGGGTCGCTTTCTT	131~156	4
DFR-2	F:GCACCGTTCGTTTATACCGC	R:GGTGGCCACCGAGAAATGTA	147~189	18
C4H	F:TCATTCGTGCCTATGTCGATACT	R:GCCTTTGTGCTCGACAGTTT	260~294	5
F3'H	F:GCCAAAGCCAAAGAAGAGGG	R:GCTTCAACAACCTTGCCATCA	385~465	8
UFGT-1	F:TCCCTCGATACATGTTGGCA	R:GCACATCTCACCACTCCTCC	288~299	8
CHS	F:GAGAAGTTCAAGCGCATGTGT	R:CCTTGGTGGCTGCTTCTTTG	484~518	6
ANR-3	F:TTGGTGGACGCTTGGTGATT	R:AGCCTCCACTCCGATCTCTT	222~258	11
ANR-5	F:CAGTAGAGTCTGCGTCACGG	R:ACTTTGGATGTGTCGCCTGT	282~311	5
FHT-4	F:CGAGTTGACACCCTCCACTC	R:ACGAAGCTTTTCCTCGGGAG	546~548	2

群体名称 Population	份数 Number	多态位点数 Np	多态位点百分率/% PPB	观测等位基因数 Na	有效等位基因数 Ne	Nei's多样性指数 H	Shannon's 信息指数 I
不涩Unastringent	19	55	59.14	1.591 4	1.196 2	0.123 8	0.200 6
稍涩Slightly astringent	54	67	72.04	1.720 4	1.195 1	0.123 1	0.201 5
较涩More astringent	36	63	67.74	1.677 4	1.201 4	0.126 5	0.205 7
涩Astringent	9	41	44.09	1.440 9	1.183 1	0.115 5	0.182 9
总计Total	118	91	97.85	1.978 5	1.198 5	0.126 3	0.209 7

Please choose a citation manager

Content to export