Abstract
In this study, we aimed to develop novel genic simple sequence repeat (eSSR) markers and to study phylogenetic relationship among Pistacia species. Transcriptome sequencing was performed in different tissues of Siirt and Atl cultivars of pistachio (Pistacia vera). A total of 37.5-Gb data were used in the assembly. The number of total contigs and unigenes was calculated as 98,831, and the length of N50 was 1,333 bp after assembly. A total of 14,308 dinucleotide, trinucleotide, tetranucleotide, pentanucleotide, and hexanucleotide SSR motifs (4–17) were detected, and the most abundant SSR repeat types were trinucleotide (29.54%), dinucleotide (24.06%), hexanucleotide (20.67%), pentanucleotide (18.88%), and tetranucleotide (6.85%), respectively. Overall 250 primer pairs were designed randomly and tested in eight Pistacia species for amplification. Of them, 233 were generated polymerase chain reaction products in at least one of the Pistacia species. A total of 55 primer pairs that had amplifications in all tested Pistacia species were used to characterize 11 P. vera cultivars and 78 wild Pistacia genotypes belonging to nine Pistacia species (P. khinjuk, P. eurycarpa, P. atlantica, P. mutica, P. integerrima, P. chinensis, P. terebinthus, P. palaestina, and P. lentiscus). A total of 434 alleles were generated from 55 polymorphic eSSR loci with an average of 7.89 alleles per locus. The mean number of effective allele was 3.40 per locus. Polymorphism information content was 0.61, whereas observed (Ho) and expected heterozygosity (He) values were 0.39 and 0.65, respectively. UPGMA (unweighted pair-group method with arithmetic averages) and STRUCTURE analysis divided 89 Pistacia genotypes into seven populations. The closest species to P. vera was P. khinjuk. P. eurycarpa was closer P. atlantica than P. khinjuk. P. atlantica–P. mutica and P. terebinthus–P. palaestina pairs of species were not clearly separated from each other, and they were suggested as the same species. The present study demonstrated that eSSR markers can be used in the characterization and phylogenetic analysis of Pistacia species and cultivars, as well as genetic linkage mapping and QTL (quantitative trait locus) analysis.
Highlights
MATERIALS AND METHODSPistacia L. genus is a member of the Anacardiaceae family that contains important species such as mango, pepper tree, and sumac (Kafkas, 2006a)
Sixteen transcriptome libraries were constructed from different tissues of Siirt and Atlı cultivars, and a total of 374,726,850 clean reads were obtained
A total of 98,831 unigenes were generated by the Trinity software, and the N50 of unigenes was computed as 1,333 bp (Supplementary Table 4)
Summary
MATERIALS AND METHODSPistacia L. genus is a member of the Anacardiaceae family that contains important species such as mango, pepper tree, and sumac (Kafkas, 2006a). The genus of Pistacia consists of 13 or more species (Gundesli et al, 2019), and Pistacia vera is believed to be the most ancestral species, whereas the other species probably derived from Zohary (1952) and Kafkas (2019). The first taxonomic study in the genus Pistacia was done morphologically by Zohary (1952). The first detailed molecular study in Pistacia was performed based on chloroplast DNA profiles by Parfitt and Badenes (1997). Microsatellites or simple sequence repeats (SSRs) and repeats of 1- to 6-nucleotide-long DNA motifs have high reproducibility, multiallelic character, and extensive tandem repeats in the whole genome (Powell et al, 1996). SSRs have advantages over other marker systems because of their codominant inheritance, suitability for automation, and well-distribution throughout eukaryotic genomes. SSRs have been widely used in genetic map construction, DNA fingerprinting, genetic diversity, quantitative trait locus (QTL) mapping, and markerassisted selection (MAS) (Dong et al, 2018; Yang et al, 2018; Zhang et al, 2019)
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