Comprehensive Characterization and Validation of Chromosome-Specific Highly Polymorphic SSR Markers From Pomegranate (Punica granatum L.) cv. Tunisia Genome

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.

Genome skimming-based simple sequence repeat (SSR) marker discovery and characterization in Grevillea robusta

“…A total of 13,335 SSRs were successfully identified, where maximum relative abundance and density were obtained for di-nucleotides (9.65 loci Mb -1 ; 140.18 bp Mb -1 ) followed by tri-, tetraand penta-nucleotides. It revealed that most of the nongenic region contains a large set of di-nucleotides repeats, which is similar to other taxon studies (Karaoglu et al 2005;Patil et al 2021;Sigang et al 2021;Zhu et al 2021). The SSR repeat analysis revealed that di-and tri-nucleotide repeats were almost ubiquitous in the genome, and these classes were further dominated by repeat motifs AG/CT and AAT/CTT, respectively.…”

Genome-Wide Survey and Development of the First Microsatellite Markers Database (AnCorDB) in Anemone coronaria L.

“…Six classes of perfect SSRs were evaluated (from mono-to hexanucleotide) for their abundance in the assembled genome. Dinucleotides were the most abundant, in accordance with what has been previously reported in literature [30][31][32][33][34][35][36][37][38][39], representing 60.2% of the identified SSRs. Trinucleotides were the second most abundant class (23.7%), followed by tetranucleotides (6.8%).…”

Chloroplast Genome Sequencing, Comparative Analysis, and Discovery of Unique Cytoplasmic Variants in Pomegranate (Punica granatum L.)

“…Pomegranate ( Punica granatum L.) is one of the oldest edible fruit crops belonging to the family Lythraceae ( Dandachi et al, 2017 ). It is believed to have originated from Iran and is widely grown in drier parts of the world, i.e., Southeast Asia, India, Iran, China, Japan, United States (California), Israel, Spain, Turkey, and Israel ( Yan et al, 2019a ; Patil et al, 2020 , 2021 ; Singh et al, 2020a ). Being a native of central Asia that is from Iran to Western Himalayas in India, later, it was domesticated to many other parts of the world ( Holland et al, 2009 ).…”