黄鳍棘鲷SNP标记开发与验证

doi:10.11978/2022108

Abstract

Abstract:

In this experiment, 'Acanthopagrus latus', an important economic fish in the southeast coast of China, was used as the experimental material. The SNPs were discovered by DNA re-sequencing in fifty A. latus, and partial SNPs were genotyped using MassARRAY^® DNA mass spectrometry. The results are presented as followed: 1) the re-sequencing of 50 wild A. latus generated a total of about 233.48 GB raw data. After filtering adapters and low-quality data, 233.43 Gb clear data were obtained. The average data size of each sample was 4.67 GB, and the average GC content was 42.85%, Q20 is above 96.56%, Q30 is above 91.1%, and the comparison rate between the clear data and the reference genome is 98.06% ~ 99.47%; 2) a total of 13843766 SNPs were discovered from 50 individuals by GATK, and 6501 high-quality SNPs were obtained after filtering; 3) thirty SNPs from the high-quality SNP were selected randomly and genotyped using MassARRAY technology. The detection rate (loci that can be genotyped) was reached at 98%. The consistency between the genome re-sequencing and the MassARRAY results was 64.83%, which indicated that the two techniques were different in detecting SNPs. In summary, a method for mining, filtering and validating SNP markers of A. latus bream has been established in this study, and the developed SNP loci can be used in evaluation of proliferation and stocking effect and genome selection breeding of A. latus in the future.

Key words: Acanthopagrus latus, Re-sequencing, SNP, MassARRAY

ZHENG Guobin, ZHAO Hongbo, HUANG Liangmin, ZHANG Jing, LIU Xiande. Discovery and verification of SNP in Acanthopagrus latus[J].Journal of Tropical Oceanography, 2023, 42(2): 78-86.

Figures/Tables 7

Tab. 1

Fig. 1

Tab. 2

The results of re-sequence data quality"

样品	原始数据/Gb	过滤数据/Gb	比对率/%	测序深度/×	Q20/%	Q30/%	GC含量/%	样品	原始数据/Gb	过滤数据/Gb	比对率/%	测序深度/×	Q20/%	Q30/%	GC含量/%
Y01	4.71	4.71	95.90	6.15	97.34	92.78	42.69	Y26	4.79	4.79	97.00	6.33	97.33	92.77	42.80
Y02	5.00	4.99	95.49	6.52	97.39	92.88	42.69	Y27	4.59	4.59	96.96	6.09	97.27	92.52	42.93
Y03	4.76	4.76	96.20	6.22	97.43	92.94	42.61	Y28	4.63	4.63	96.76	6.10	96.56	91.10	42.78
Y04	4.55	4.55	96.83	6.03	97.49	93.13	42.73	Y29	4.74	4.74	96.74	6.27	97.34	92.84	42.83
Y05	4.58	4.58	96.38	6.06	97.43	93.03	42.76	Y30	4.48	4.48	96.96	5.96	97.54	93.17	42.66
Y06	4.46	4.46	96.46	5.92	97.44	93.02	42.69	Y31	4.85	4.85	97.2	6.41	97.47	93.06	42.66
Y07	4.71	4.71	97.03	6.25	97.45	93.06	42.76	Y32	4.17	4.17	97.10	5.59	97.26	92.60	42.75
Y08	4.79	4.79	96.69	6.32	97.21	92.52	42.80	Y33	4.98	4.98	97.27	6.55	97.27	92.68	42.99
Y09	4.86	4.86	97.36	6.43	97.34	92.82	43.18	Y34	3.69	3.69	96.8	5.00	97.29	92.64	42.96
Y10	4.78	4.77	96.60	6.30	97.19	92.46	42.87	Y35	4.92	4.92	97.33	6.48	97.12	92.31	43.03
Y11	4.43	4.43	96.56	5.88	96.97	92.00	42.83	Y36	4.86	4.86	97.64	6.41	97.23	92.58	42.95
Y12	4.59	4.59	96.60	6.06	97.09	92.23	42.95	Y37	4.88	4.88	97.32	6.45	97.04	92.13	42.79
Y13	4.63	4.63	96.87	6.13	97.26	92.64	42.78	Y38	4.84	4.84	96.84	6.35	97.25	92.59	42.74
Y14	4.82	4.82	96.55	6.32	97.08	92.24	42.71	Y39	4.59	4.59	96.43	6.07	97.28	92.65	42.85
Y15	4.63	4.63	96.57	6.12	97.22	92.49	42.77	Y40	4.23	4.23	97.54	5.66	97.36	92.82	42.95
Y16	4.70	4.69	96.80	6.20	97.24	92.61	42.97	Y41	4.73	4.73	97.65	6.26	97.49	93.13	43.01
Y17	4.93	4.93	96.95	6.50	97.17	92.43	42.91	Y42	4.37	4.37	96.58	5.77	97.26	92.61	43.08
Y18	4.33	4.33	97.05	5.79	97.10	92.22	42.81	Y43	4.90	4.90	97.08	6.45	97.42	92.99	43.01
Y19	4.85	4.85	96.91	6.40	97.25	92.59	42.95	Y44	4.82	4.81	97.36	6.36	97.49	93.09	42.99
Y20	4.52	4.52	96.45	5.96	97.33	92.76	42.75	Y45	4.95	4.95	97.14	6.52	97.50	93.16	43.02
Y21	4.79	4.79	97.00	6.32	97.20	92.46	42.73	Y46	4.61	4.61	97.09	5.85	97.46	93.06	42.68
Y22	4.84	4.84	97.00	6.36	97.54	93.21	42.54	Y47	4.75	4.75	96.72	6.27	97.42	92.98	42.86
Y23	4.71	4.71	96.41	6.17	96.71	91.43	42.72	Y48	4.50	4.50	96.85	5.96	97.44	93.02	42.90
Y24	4.58	4.58	96.84	6.08	97.25	92.56	42.75	Y49	4.42	4.42	97.43	5.90	97.17	92.46	43.33
Y25	4.75	4.75	97.22	6.27	97.33	92.74	42.80	Y50	4.88	4.88	97.36	6.44	97.38	92.88	43.03

Tab. 2

Tab. 3

Fig. 2

Tab. 4

Tab. 5

Genotyping comparison between genome re-sequencing and MassARRAY"

SNP	染色体名称	位置	MassARRAY分型检测				全基因组重测序
SNP	染色体名称	位置	检测结果	检出率/%	MF	MAF	检测结果	检出率/%	MF	MAF
Shq_178_1	NC_051039.1	33527645	CC、CG	85	0.85	0.15	CC、CG	95	0.74	0.26
Shq_178_2	NC_051040.1	32016498	TT	100	1.00	0	TT、TG	100	0.88	0.13
Shq_178_3	NC_051041.1	20806265	TT、TG	100	0.95	0.05	TT、TG	100	0.75	0.25
Shq_178_6	NC_051042.1	30329	AA、AG	90	0.69	0.31	AA、AG	95	0.63	0.37
Shq_178_8	NC_051042.1	7311706	TT、TC	100	0.95	0.05	TT、TC	100	0.95	0.05
Shq_178_19	NC_051048.1	22404460	AA	95	1.00	0	AA、AG	90	0.92	0.08
Shq_178_26	NC_051050.1	34885	CC	95	1.00	0	CC、CA	95	0.92	0.08
Shq_178_39	NC_051050.1	38757	GG	100	1.00	0	GG、GA	90	0.94	0.06
Shq_178_40	NC_051050.1	42037	TT	100	1.00	0	TT、TC	95	0.87	0.13
Shq_178_48	NC_051050.1	59154	TT、TC	85	0.91	0.09	TT、TC	100	0.65	0.35
Shq_178_60	NC_051050.1	74750	GG	100	1.00	0	GG、GA	90	0.94	0.06
Shq_178_63	NC_051050.1	75488	GG	100	1.00	0	GG、GC	95	0.95	0.05
Shq_178_65	NC_051050.1	76081	AA	100	1.00	0	AA、AC	85	0.88	0.12
Shq_178_70	NC_051050.1	87063	GG	100	1.00	0	GG、GA	90	0.89	0.11
Shq_178_72	NC_051050.1	93481	CC、CT	100	0.98	0.03	CC、CT	100	0.95	0.05
Shq_178_73	NC_051050.1	96282	CC	100	1.00	0	CC、CA	95	0.82	0.18
Shq_178_77	NC_051050.1	101791	CC	100	1.00	0	CC、CT	80	0.94	0.06
Shq_178_86	NC_051050.1	106638	GG、GA	100	0.97	0.03	GG、GA	80	0.97	0.03
Shq_178_92	NC_051050.1	128900	AA	100	1.00	0	AA、AG	100	0.93	0.08
Shq_178_96	NC_051050.1	142395	GG	100	1.00	0	GG、GA	90	0.92	0.08
Shq_178_104	NC_051052.1	1588997	AA	100	1.00	0	AA、AC	100	0.75	0.25
Shq_178_112	NC_051053.1	1538	GG	100	1.00	0	AA、AG、GG	95	0.53	0.47
Shq_178_138	NC_051053.1	8936396	AA	100	1.00	0	AA、AG	95	0.76	0.24
Shq_178_149	NC_051056.1	31720087	CC、CT	100	0.95	0.05	CC、CT	90	0.86	0.14
Shq_178_150	NC_051056.1	31723482	TT	100	1.00	0	TT、TC	95	0.89	0.11
Shq_178_157	NC_051058.1	24478462	AA、AG	100	0.95	0.05	AA、AC	100	0.95	0.05
Shq_178_158	NC_051058.1	24478507	CC、CT	95	0.97	0.03	CC、CT	100	0.95	0.05
Shq_178_171	NC_051060.1	26149536	GG	100	1.00	0	GG、GC	100	0.58	0.43
Shq_178_172	NC_051061.1	336649	TT	100	1.00	0	TT、TG	100	0.55	0.45
Shq_178_173	NC_051061.1	22105514	GG	100	1.00	0	GG、GA	100	0.55	0.45

Tab. 5

References 30

[1]	江兴龙, 黄永春, 黄良敏, 等, 2013. 厦门湾黄鳍鲷增殖放流效果的评估[J]. 集美大学学报(自然科学版), 18(3): 161-166.
	JIANG XINGLONG, HUANG YONGCHUN, HUANG LIANGMIN, et al, 2013. An evaluation on the effect of Sparus latus enhancement ＆ release in Xiamen Bay[J]. Journal of Jimei University (Natural Science Edition), 18(3): 161-166. (in Chinese with English abstract)
[2]	吴利娜, 张凝鋆, 孙松, 等, 2021. 微卫星分子标记技术在大黄鱼增殖放流效果评估中的应用[J]. 中国水产科学, 28(9): 1100-1108.
	WU LINA, ZHANG NINGJUN, SUN SONG, et al, 2021. Application of microsatellite markers for evaluating the effect of restocking enhancement in Larimichthys crocea[J]. Chinese Fishery Sciences, 28(9): 1100-1108. (in Chinese with English abstract)
[3]	杨习文, 刘熠, 薛向平, 等, 2020. 基于微卫星标记的长江江苏段鲢(Hypophthalmichthys molitrix)增殖放流资源贡献率的评估[J]. 湖泊科学, 32(4): 1154-1164.
	YANG XIWEN, LIU YI, XUE XIANGPING, et al, 2020. Resource contribution rate assessment of stock enhancement of silver carp, Hypophthalmichthy smolitrix in Jiangsu section of the Yangtze River based on microsatellite markers[J]. Lake Science, 32(4): 1154-1164. (in Chinese with English abstract)
[4]	赵雨, 2021. 基于微卫星标记的日本对虾增殖放流效果评价及群体遗传学研究[D]. 天津农学院.
	ZHAO YU, 2021. Study on stock enhancement and genetics of Penaeus japonicus based on microsatellite markers[D]. Tianjin Agricultural University. (in Chinese with English abstract)
[5]	朱克诚, 宋岭, 刘宝锁, 等, 2020. 黄鳍棘鲷家系亲缘关系鉴定[J]. 水产学报, 44(3): 351-357.
	ZHU KECHENG, SONG LING, LIU BAOSUO, et al, 2020. Establishment of parentage determination in yellowfin seabream (Acanthopagrus latus)[J]. Journal of Fisheries, 44(3): 351-357. (in Chinese with English abstract)
[6]	BEACHAM T D, WALLACE C, JONSEN K, et al, 2018. Comparison of coded-wire tagging with parentage-based tagging and genetic stock identification in a large-scale coho salmon fisheries application in British Columbia, Canada[J]. Evolutionary Applications, 12(2): 230-254. doi: 10.1111/eva.12711
[7]	BUDOWLE B, 2004. SNP typing strategies[J]. Forensic Science International, 146: 139-142.
[8]	CHEN SHIFU, ZHOU YANQING, CHEN YARU, et al, 2018. Fastp: an unltra-fast all-in-one FASTQ preprocessor[J]. Bioinformatics, 34(17): 884-890. doi: 10.1093/bioinformatics/btx692
[9]	DING WEIDONG, ZHANG XINHUI, ZHAO XIAOMENG, et al, 2021. A chromosome-level genome assembly of the Mandarin fish(Siniperca chuatsi)[J]. Frontiers in Genetics, 12: 671650. doi: 10.3389/fgene.2021.671650
[10]	ELLIS J A, ONG B, 2017. The MassARRAY^® system for targeted SNP genotyping[M]// Genotyping. New York, NY: Humana Press: 77-94.
[11]	ETTER P D, BASSHAM S, HOHENLOHE P A, et al, 2012. SNP discovery and genotyping for evolutionary genetics using RAD sequencing[M]// Molecular methods for evolutionary genetics. Humana Press: 157-178.
[12]	GARCÍA-FERNÁNDEZ C, SÁNCHEZ J A, BLANCO G, 2018. SNP-haplotypes: An accurate approach for parentage and relatedness inference in gilthead sea bream (Sparus aurata)[J]. Aquaculture, 495: 582-591. doi: 10.1016/j.aquaculture.2018.06.019
[13]	JURINKE C, VAN DEN BOOM D, CANTOR C R, et al, 2002. The use of MassARRAY technology for high throughput genotyping[J]. Chip Technology, 77: 57-74.
[14]	KUMAR S, BANKS T W, CLOUTIER S, 2012. SNP discovery through next-generation sequencing and its applications[J]. International journal of Plant Genomics.
[15]	LI HENG, 2013. Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM[J]. ArXiv, 1303: 3997.
[16]	LI HENG, HANDSAKER B, WYSOKER A, et al, 2009. The sequence alignment/map format and SAMtools[J]. Bioinformatics, 25(16): 2078-2079. doi: 10.1093/bioinformatics/btp352 pmid: 19505943
[17]	LIU SIXIN, PALTI Y, GAO GUANGTU, et al, 2016. Development and validation of a SNP panel for parentage assignment in rainbow trout[J]. Aquaculture, 452: 178-182. doi: 10.1016/j.aquaculture.2015.11.001
[18]	MCKENNA A, HANNA M, BANKS E, et al, 2010. The genome analysis toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data[J]. Genome Res, 20(9): 1297-1303. doi: 10.1101/gr.107524.110 pmid: 20644199
[19]	MEHTA B, DANIEL R, PHILLIPS C, et al, 2017. Forensically relevant SNaPshot^® assays for human DNA SNP analysis: a review[J]. International Journal of Legal Medicine, 131(1): 21-37. doi: 10.1007/s00414-016-1490-5
[20]	MONTES I, CONKLIN D, ALBAINA A, et al, 2013. SNP discovery in european anchovy (Engraulis encrasicolus, L) by high-throughput transcriptome and genome sequencing[J]. PLoS One, 8(8): e70051. doi: 10.1371/journal.pone.0070051
[21]	OETH P, PARK C, KOSMAN D, et al, 2005. iPLEX assay: Increased plexing efficiency and flexibility for MassArray^® system through single base primer extension with mass-modified terminators[J]. Sequenom Application Note, 27.
[22]	OHNISHI Y, TANAKA T, OZAKI K, et al, 2001. A high-throughput SNP typing system for genome-wide association studies[J]. Journal of Human Genetics, 46(8): 471-477. doi: 10.1007/s100380170047 pmid: 11501945
[23]	RODI C P, DARNHOFER-PATEL B, STANSSENS P, et al, 2002. A strategy for the rapid discovery of disease markers using the MassARRAY system[J]. Biotechniques, Suppl: 62-66, 68-69.
[24]	SEO S B, KING J L, WARSHAUER D H, et al, 2013. Single nucleotide polymorphism typing with massively parallel sequencing for human identification[J]. International Journal of Legal Medicine, 127(6): 1079-1086. doi: 10.1007/s00414-013-0879-7 pmid: 23736940
[25]	SUN XIAOWEN, LIU DONGYUAN, ZHANG XIAOFENG, et al, 2013. SLAF-seq: an efficient method of large-scale de novo SNP discovery and genotyping using high-throughput sequencing[J]. PloS One, 8(3): e58700. doi: 10.1371/journal.pone.0058700
[26]	VERA M, ALVAREZ-DIOS J A, FERNANDEZ C, et al, 2013. Development and validation of single nucleotide polymorphisms (SNPs) markers from two transcriptome 454-runs of turbot (Scophthalmus maximus) using high-throughput genotyping[J]. International Journal of Molecular Sciences, 14(3): 5694-5711. doi: 10.3390/ijms14035694 pmid: 23481633
[27]	WANG YONGPING, GUO XIMING, 2008. ITS Length polymorphism in oysters and its use in species identification[J]. Journal of Shellfish Research, 27(3): 489-493. doi: 10.2983/0730-8000(2008)27[489:ILPIOA]2.0.CO;2
[28]	YÁÑEZ J M, NASWA S, LÓPEZ M E, et al, 2016. Genomewide single nucleotide polymorphism discovery in Atlantic salmon (Salmo salar): validation in wild and farmed American and European populations[J]. Molecular Ecology Resources, 16(4): 1002-1011. doi: 10.1111/1755-0998.12503 pmid: 26849107
[29]	YÁÑEZ J M, YOSHIDA G, BARRIA A, et al, 2020. High-throughput single nucleotide polymorphism (SNP) discovery and validation through whole-genome resequencing in Nile tilapia (Oreochromis niloticus)[J]. Marine Biotechnology, 22(1): 109-117. doi: 10.1007/s10126-019-09935-5 pmid: 31938972
[30]	ZHAO H, Li C, HARGROVE J S, et al, 2018. SNP marker panels for parentage assignment and traceability in the Florida bass (Micropterus floridanus)[J]. Aquaculture, 485: 30-38. doi: 10.1016/j.aquaculture.2017.11.014

取样地	数量/尾	体长/cm	体重/g	体厚/cm
厦门近海海区	37	20.52 ± 0.10	190.15 ± 5.92	7.27 ± 0.05
漳州东山岛海区	13	21.23 ± 0.29	212.53 ± 6.19	7.52 ± 0.08

染色体名称	SNP初始数	质控过滤后SNP数	属性过滤后SNP数	高质量过滤后SNP数
NC_051039.1	692976	632996	144274	364
NC_051040.1	646881	595158	134036	221
NC_051041.1	566757	522152	117157	325
NC_051042.1	690338	640940	142536	172
NC_051043.1	606086	565937	129160	157
NC_051044.1	697405	638444	144000	265
NC_051045.1	589449	548080	124463	366
NC_051046.1	615425	574376	130360	228
NC_051047.1	622596	577032	131271	203
NC_051048.1	476295	405575	97778	820
NC_051049.1	629215	586030	132799	263
NC_051050.1	574454	532521	120384	237
NC_051051.1	567923	530390	119320	139
NC_051052.1	462308	429823	94073	109
NC_051053.1	538049	501748	110871	172
NC_051054.1	535520	495987	112649	173
NC_051055.1	613575	567469	128275	221
NC_051056.1	669814	618189	137376	256
NC_051057.1	520747	477109	104765	187
NC_051058.1	509767	475642	106603	97
NC_051059.1	583650	535046	121219	370
NC_051060.1	563226	523705	117774	169
NC_051061.1	518795	479194	110565	299
NC_051062.1	329951	298031	67740	688
未知序列	22564	0	0	0
合计	13843766	12751574	2879448	6501

多态性参数	最大值	最小值	平均值
观测杂合度(H_o)	0.966	0.034	0.499
期望杂合度(H_e)	0.600	0.034	0.376
多态信息含量(PIC)	0.375	0.033	0.279

Discovery and verification of SNP in Acanthopagrus latus

RichHTML

PDF (PC)

Like

Knowledge

Abstract

Cite this article

share this article

Figures/Tables 7

References 30

Related Articles 2

Metrics

Comments

Recommended 0

[1]	HUANG Jing, PAN Xiaolan, XU Meng, LIU Wenguang, ZHANG Hua, HE Maoxian. Morphological and SNP markers for analysis of genetic structure of hybrid progeny and their parental populations of Pinctada fucata martensii [J]. Journal of Tropical Oceanography, 2019, 38(6): 80-89.
[2]	Jialin AI, Zhimin LI, Jianyong LIU, Yuchun SHEN. Single nucleotide polymorphisms of myostation gene and its association with growth traits in Haliotis diversicolor supertexta [J]. Journal of Tropical Oceanography, 2019, 38(2): 78-85.