Abstract
BackgroundPistachio (Pistacia vera L.) is a dioecious species that has a long juvenility period. Therefore, development of marker-assisted selection (MAS) techniques would greatly facilitate pistachio cultivar-breeding programs. The sex determination mechanism is presently unknown in pistachio. The generation of sex-linked markers is likely to reduce time, labor, and costs associated with breeding programs, and will help to clarify the sex determination system in pistachio.ResultsRestriction site-associated DNA (RAD) markers were used to identify sex-linked markers and to elucidate the sex determination system in pistachio. Eight male and eight female F1 progenies from a Pistacia vera L. Siirt × Bağyolu cross, along with the parents, were subjected to RAD sequencing in two lanes of a Hi-Seq 2000 sequencing platform. This generated 449 million reads, comprising approximately 37.7 Gb of sequences. There were 33,757 polymorphic single nucleotide polymorphism (SNP) loci between the parents. Thirty-eight of these, from 28 RAD reads, were detected as putative sex-associated loci in pistachio. Validation was performed by SNaPshot analysis in 42 mature F1 progenies and in 124 cultivars and genotypes in a germplasm collection. Eight loci could distinguish sex with 100% accuracy in pistachio. To ascertain cost-effective application of markers in a breeding program, high-resolution melting (HRM) analysis was performed; four markers were found to perfectly separate sexes in pistachio. Because of the female heterogamety in all candidate SNP loci, we report for the first time that pistachio has a ZZ/ZW sex determination system. As the reported female-to-male segregation ratio is 1:1 in all known segregating populations and there is no previous report of super-female genotypes or female heteromorphic chromosomes in pistachio, it appears that the WW genotype is not viable.ConclusionSex-linked SNP markers were identified and validated in a large germplasm and proved their suitability for MAS in pistachio. HRM analysis successfully validated the sex-linked markers for MAS. For the first time in dioecious pistachio, a female heterogamety ZW/ZZ sex determination system is suggested.
Highlights
Pistachio (Pistacia vera L.) is a dioecious species that has a long juvenility period
450,721,882 reads, comprising approximately 36,959,194,324 nucleotide sequences were obtained following cleaning of multiplex identifier (MID) sequences. 18.49 Gb of data was obtained from female plants and 18.47 Gb from male plants
Thirty-eight putative sex-linked markers were detected from 28 reads following restriction site-associated DNA sequencing (RADseq)
Summary
Pistachio (Pistacia vera L.) is a dioecious species that has a long juvenility period. Development of marker-assisted selection (MAS) techniques would greatly facilitate pistachio cultivar-breeding programs. The sex determination mechanism is presently unknown in pistachio. The generation of sex-linked markers is likely to reduce time, labor, and costs associated with breeding programs, and will help to clarify the sex determination system in pistachio. Pistacia vera is the only commercially important species, and is believed to be the most ancestral species, other species are probably its derivatives [3]. Numerous DNA-based molecular marker systems have been developed for marker-assisted selection (MAS) in breeding programs. Next-generation sequencing (NGS) technology provides an effective tool for MAS through generating huge number of DNA markers within a short period.
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