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Genome Wide Characterization of Simple Sequence Repeats in Watermelon Genome and Their Application in Comparative Mapping and Genetic Diversity Analysis

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Genome Wide Characterization of Simple Sequence Repeats in Watermelon Genome and Their Application in Comparative Mapping and Genetic Diversity Analysis

Huayu Zhu et al. BMC Genomics.

Abstract

Background: Microsatellite markers are one of the most informative and versatile DNA-based markers used in plant genetic research, but their development has traditionally been difficult and costly. The whole genome sequencing with next-generation sequencing (NGS) technologies provides large amounts of sequence data to develop numerous microsatellite markers at whole genome scale. SSR markers have great advantage in cross-species comparisons and allow investigation of karyotype and genome evolution through highly efficient computation approaches such as in silico PCR. Here we described genome wide development and characterization of SSR markers in the watermelon (Citrullus lanatus) genome, which were then use in comparative analysis with two other important crop species in the Cucurbitaceae family: cucumber (Cucumis sativus L.) and melon (Cucumis melo L.). We further applied these markers in evaluating the genetic diversity and population structure in watermelon germplasm collections.

Results: A total of 39,523 microsatellite loci were identified from the watermelon draft genome with an overall density of 111 SSRs/Mbp, and 32,869 SSR primers were designed with suitable flanking sequences. The dinucleotide SSRs were the most common type representing 34.09 % of the total SSR loci and the AT-rich motifs were the most abundant in all nucleotide repeat types. In silico PCR analysis identified 832 and 925 SSR markers with each having a single amplicon in the cucumber and melon draft genome, respectively. Comparative analysis with these cross-species SSR markers revealed complicated mosaic patterns of syntenic blocks among the genomes of three species. In addition, genetic diversity analysis of 134 watermelon accessions with 32 highly informative SSR loci placed these lines into two groups with all accessions of C.lanatus var. citorides and three accessions of C. colocynthis clustered in one group and all accessions of C. lanatus var. lanatus and the remaining accessions of C. colocynthis clustered in another group. Furthermore, structure analysis was consistent with the dendrogram indicating the 134 watermelon accessions were classified into two populations.

Conclusion: The large number of genome wide SSR markers developed herein from the watermelon genome provides a valuable resource for genetic map construction, QTL exploration, map-based gene cloning and marker-assisted selection in watermelon which has a very narrow genetic base and extremely low polymorphism among cultivated lines. Furthermore, the cross-species transferable SSR markers identified herein should also have practical uses in many applications in species of Cucurbitaceae family whose whole genome sequences are not yet available.

Keywords: Comparative genomics; Cucurbits; Genetic diversity; SSR; Synteny; Watermelon.

Figures

Fig. 1
Fig. 1
Distribution of SSR motif repeat numbers and relative frequency in watermelon genome. The vertical axis shows the abundance of microsatellites that have different motif repeat numbers (from 3 to > 20), which are discriminated by legends of different colours
Fig. 2
Fig. 2
The distribution of SSR repeat types on each chromosome in watermelon. The vertical axis shows the number of microsatellites from dinucleotide to octonucleotide which are discriminated by different colours. The horizontal axis show different chromosomes of watermelon and chr0 represent all the chromosomal unanchored scaffolds
Fig. 3
Fig. 3
Syntenic relationships of watermelon with cucumber (a) and melon (b) chromosomes. Chromosome synteny between watermelon and cucumber is based on 821 cross-species markers (A); synteny between watermelon and melon is based on 850 cross-species markers. W1-W11 represent watermelon eleven chromosomes, C1-C7 represent cucumber seven chromosomes and I-XII represent melon twelve chromsomes. Syntenic blocks are connected by with the same colour lines from watermelon chromosomes
Fig. 4
Fig. 4
A syntenic block view of cucumber and melon chromosomes composted of watermelon chromosomes. The different colours represent the eleven chromosomes of watermelon. The mosaic colour pattern of cucumber and melon chromosomes indicated seven cucumber chromosomes and twelve melon chromosomes composed of syntenic blocks from different watermelon chromosomes
Fig. 5
Fig. 5
Comparative pachytene FISH analysis of cucumber (C7), melon (I) and watermelon (W2 and W9). Fourteen fosmid probes identified on cucumber chromosome C7 and melon chromosome I in a previous study (Yang et al, [12]) were used to detect their location in watermelon for verifying the results by comparative mapping. CEN indicates the putative centromere location
Fig. 6
Fig. 6
The UPGMA phylogenetic tree of the 134 accessions. The phylogenetic analysis showed 134 accessions were classified into two groups: group I and II. The colour branches represent the accessions collected from different continents. The number 1–134 represent waermelon accessions W1 to W134 in Additional file 2: Table S1. Among them, 1–5 belong to C. colocynthis, 6–17 belong to C. lanatus var. citroides and 18–134 belong to C. lanatus var. lanatus
Fig. 7
Fig. 7
Population structure of 134 accessions in watermelon by Model-based analysis. Scale of Y axis represents the percent of genetic components, and the X axis represents the different watermelon accession. The colour dots in the top of these bar plots represent the origin of these accessions, and the latin number (I and II) corresponds to the predefined phylogenetic tree in Fig. 6

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