Scientia Agricultura Sinica ›› 2023, Vol. 56 ›› Issue (8): 1547-1560.doi: 10.3864/j.issn.0578-1752.2023.08.010

• HORTICULTURE • Previous Articles     Next Articles

Population Genomic Structure of Pomelo Germplasm and Fruit Acidity Associated Genes Identification by Genotyping-by-Sequencing Technology

JIANG Dong1(), WANG Xu1, LI RenJing1, ZHAO XiaoDong2, DAI XiangSheng2, LIU ZhengWei3   

  1. 1 Citrus Research Institute, Southwest University, Chongqing 400712
    2 Jinggangshan Agricultural Science and Technology Park Management Committee, Ji’an 343016, Jiangxi
    3 Jianggangshan University, Ji’an 343016, Jiangxi
  • Received:2022-05-25 Accepted:2022-10-08 Online:2023-04-16 Published:2023-04-23

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Abstract:

【Objective】 To reveal the phylogeny, population genetic structure and diversity level of pomelo (Citrus grandis (L.) Osbeck) germplasm, and to efficiently utilize them to explore genes related to important fruit quality traits, this research provided an insight into the population genetic structure and phylogeny of pomelo germplasm and facilitated the pomelo varieties innovation.【Method】 A total of 282 pomelo accessions including landraces from different geographical regions and hybrid offspring of kiyomi tangor and pomelo were contained in this study. GBS library was constructed with genomic DNAs digested by EcoR I restriction endonuclease and sequenced on Illumina HiSeq PE150 platform, the clean short reads were then mapped to pomelo reference genome by BWA, and SNPs were called out with SAMTOOLS pipeline. Based on 121 726 SNPs genotyping data, principal component analysis (PCA) and population genetic structure analysis were carried out and phylogenetic trees were constructed with Neighbor-joining method. Furthermore, two sub-populations containing high-acid accessions (32) and low-acid accessions (23) were used to identify candidate genes related to fruit acidity by Fst and XP-CLR selective sweeping analysis. Meanwhile, the genotype data of 282 pomelo accessions and the phenotypic data of titratable acid content in fruit were used for GWAS.【Result】 A total of 201.66 Gb original reads were generated from 282 pomelo germplasm by GBS approach, in average each sample produced 0.72 Gb reads. After the screening conditions of sequencing depth of dp5, the miss less than 0.2 and minor alleles frequency (MAF)>0.01, and a total of 121 726 SNPs were selected out for subsequent analysis. The PCA, structure and phylogenetic analysis all supported that the 282 pomelo germplasm could be divided into 6 subgroups, among which pomelo and kiyomi hybrid population, grapefruits and other pomelo hybrid populations could be obviously different from true-to-type pomelo populations, pomelos originated from different geographical region displayed unique genetic feature. The pomelo germplasm from Thailand and Vietnam formed a relatively unique group different from other domestic groups, such as ShaTian pomelo, Wen Dan pomelo, and Dian Jiang pomelo in China. The genetic introgression from Vietnam pomelos were exhibited in most pomelo germplasm in southern China, suggested that Vietnam was the origin center for pomelo. In addition, some pomelo germplasm with unknown origin have been identified accurately by GBS technology. This study showed that different geographical distribution and artificial selection pressure had great effect on the genomic composition of pomelo. Besides, Fst, XP-CLR selective sweeping analysis revealed a strong selection signal region on chromosome 7 contained genes annotated as dihydrolipoyl transacetylase (DLT-E2) of pyruvate dehydrogenase complex (PDC) and aluminum-activated malate transporter (ALMT9), which involved in the synthesis and transportation of citric acid. In additional GWAS genome-wide association analysis identified another region on chromosome 2, which was also highly associated with fruit acidity. 【Conclusion】GBS technology provided reliable and efficient method for studying the phylogeny and evolution of pomelo. The study showed that artificial cross breeding, long-term artificial selection, geography isolation and domestication were the major driving forces for the formation of different types of pomelo germplasm. In addition, it clearly showed that Southeast Asian was primary center for pomelo origin and China mainland was secondary evolutionary center. Several candidate genes related to citric acid content in pomelo fruits were identified by Fst, XP-CLR selective sweeping and GWAS. This study provided important gene resources for the further genetic improvement and breeding of pomelo fruits.

Key words: GBS, pomelo phylogeny, genes related to fruit acidity, GWAS

Fig. 1

The principal component analysis of 282 pomelos accessions with 121726 SNPs The left image is based on the 1st and 2nd principal components, and the right image is based on the 1st and 3rd principal components. The red circles represent the sample from hybrid of Kiyomi and pomelo, the black circles represent the true-to-type pomelo sample, and the blue and other colored circles represent the grapefruit sample"

Fig. 2

Population genetic structure plot of 282 pomelo accessions determined by K=3 and K=6 using 121726 SNPs"

Table 2

The genetic diversity of 6 sub-population of pomelo germplasm"

亚群
Sub_population		 π值范围
Range of π		 平均π值
Average value of π		 平均Tajima’s D
Average value of Tajima’s D
沙田柚类 Shatian pomelo group 0.04—0.51 0.129 -0.15
文旦柚类 Wendan pomelo group 0.02—0.51 0.130 0.41
垫江柚类 Dianjiang pomelo group 0.03—0.51 0.134 0.45
越南泰国柚类 Vietnam and Thailand pomelo group 0.01—0.50 0.155 0.17
‘清见’杂交柚类 Pomelo and Kiyomi crossing group 0.01—0.50 0.233 1.12
葡萄柚类 Grapefruit group 0.03—0.51 0.259 0.92

Fig. 3

NJ clustering dendrogram generated using 121726 SNPs of 282 pomelo accessions"

Fig. 4

Genome-wide Fst (A) and XP-CLR selective sweep analysis (B) using two high-acid and low-acid pomelo populations The corresponding maximum position of Fst and XP-CLR is on the same location of chromosome 7"

Fig. 5

The Manhattan (A) and QQ plot (B) display significant candidate genes loci related to fruit acidity identified by genome-wide association analysis (GWAS) on 282 pomelo accessions"

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