Abstract
The simple sequence repeat (SSR) survey of ‘Tunisia’ genome (296.85 Mb) identified a total of 365,279 perfect SSRs spanning eight chromosomes, with a mean marker density of 1,230.6 SSRs/Mb. We found a positive trend in chromosome length and the SSR abundance as marker density enhanced with a shorter chromosome length. The highest number of SSRs (60,708) was mined from chromosome 1 (55.56 Mb), whereas the highest marker density (1,294.62 SSRs/Mb) was recorded for the shortest chromosome 8 (27.99 Mb). Furthermore, we categorized all SSR motifs into three major classes based on their tract lengths. Across the eight chromosomes, the class III had maximum number of SSR motifs (301,684, 82.59%), followed by the class II (31,056, 8.50%) and the class I (5,003, 1.37%). Examination of the distribution of SSR motif types within a chromosome suggested the abundance of hexanucleotide repeats in each chromosome followed by dinucleotides, and these results are consistent with ‘Tunisia’ genome features as a whole. Concerning major repeat types, AT/AG was the most frequent (14.16%), followed by AAAAAT/AAAAAG (7.89%), A/C (7.54%), AAT/AAG (5.23%), AAAT/AAAG (4.37%), and AAAAT/AAAAG (1.2%) types. We designed and validated a total of 3,839 class I SSRs in the ‘Tunisia’ genome through electronic polymerase chain reaction (ePCR) and found 1,165 (30.34%) SSRs producing a single amplicon. Then, we selected 906 highly variable SSRs (> 40 nt) from the ePCR-verified class I SSRs and in silico validated across multiple draft genomes of pomegranate, which provided us a subset of 265 highly polymorphic SSRs. Of these, 235 primers were validated on six pomegranate genotypes through wet-lab experiment. We found 221 (94%) polymorphic SSRs on six genotypes, and 187 of these SSRs had ≥ 0.5 PIC values. The utility of the developed SSRs was demonstrated by analyzing genetic diversity of 30 pomegranate genotypes using 16 HvSSRs spanning eight pomegranate chromosomes. In summary, we developed a comprehensive set of highly polymorphic genome-wide SSRs. These chromosome-specific SSRs will serve as a powerful genomic tool to leverage future genetic studies, germplasm management, and genomics-assisted breeding in pomegranate.
Highlights
Pomegranate (Punica granatum L.) is an economically important perennial fruit crop (Patil et al, 2020c)
The simple sequence repeat (SSR) were surveyed in the available genome assembly of pomegranate cultivar Tunisia for the presence of monoto hexanucleotides having a tract length of ≥ 12 bp
The study provides a large number of chromosome-specific hypervariable (3,839) and highly variable SSRs (> 40 nt, 906) for genotyping applications in pomegranate
Summary
Pomegranate (Punica granatum L.) is an economically important perennial fruit crop (Patil et al, 2020c). The popularity of pomegranate worldwide stems from its nutritional values, specific organoleptic characteristics, and a variety of health benefits (Filannino et al, 2013; Teixeira da Silva et al, 2013) This crop has originated from Iran and is widely cultivated in drier parts of Southeast Asia, Iran, China, Japan, the United States (California), West Indies, tropical America, and India (Holland and Bar-Ya’akov, 2014). It is considered an excellent fruit crop for arid zones owing to its drought tolerance. India stands first in pomegranate cultivation with an area of 2.46 hundred thousand hectare, and production and productivity of 27.91 hundred thousand metric tonnes and 12 tonnes/ha, respectively (NHB, 2018)
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