An analytical toolkit for polyploid willow discrimination

Abstract

Polyploid breeding is an important means for creating elite willow cultivars, and therefore provokes an active demand for discriminating the ploidy levels of natural willow stands. In this study, we established an analytical toolkit for polyploid willow identification by combining molecular markers and flow cytometry (FCM). A total of 10 single-copy fully informative SSRs were chosen for marker-aided selection based on a segregation test with a full-sib willow pedigree and a mutability test with a collection of natural willow stands. Aided by these molecular markers, we performed polyploid selection in two tree species and two shrub species of the genus Salix. The ploidy levels of the investigated samples were further examined using a flow cytometer. It was previously shown that results from marker-aided selection were consistent with those from FCM measurements. Based on ploidy level assessment in different willow species, it was found that tree willows were dominantly tetraploid, whereas shrub willows were most frequently diploid. With this analytical toolkit, polyploids can be rapidly screened from a large number of natural stands; thereafter, the exact ploidy levels of the polyploid candidates can be efficiently confirmed by FCM. This analytical toolkit will greatly enhance polyploid breeding programs for willows.

Relationships and genome evolution of polyploid Salix species revealed by RAD sequencing data

“…In the Chamaetia/Vetrix clade, which comprises about three quarters of all Salix species, more than one third of all species with known ploidy levels are tetra-, hexa- or octoploids. An analytical toolkit for the discrimination of polyploid willows has been established, based on the combination of SSR markers and flow cytometry, but was only used to detect the ploidy level for breeding purposes (Guo et al 2016). In subgenus Salix , the tetraploid S. alba – S. fragilis complex has been analyzed with AFLP markers and revealed an allotetraploid nature of the species (Barcaccia et al 2014), but in the Chamaetia/Vetrix clade the polyploid species have never been analyzed with molecular markers.…”

Genome-wide investigation of the AP2/ERF superfamily and their expression under salt stress in Chinese willow (Salix matsudana)

“…Salix is one of the few woody plants with a large number of polyploid taxa, in Salix matsudana, both tetraploid and diploid individuals have been observed. [25]. In our experiment, we sequenced the genome of tetraploid Salix matsudana.…”

Development of novel EST-SSR markers for ploidy identification based on de novo transcriptome assembly for Misgurnus anguillicaudatus

“…There are some important considerations that are necessary when developing SSR markers for ploidy identification. Importantly, the efficiency of SSR-assisted ploidy identification depends on the variability of amplified loci [ 40 ]. We measured the PIC value, which provides an estimate of the variability and discriminatory power of a locus by taking into account, not only the number of alleles that are expressed, but also the relative frequencies of those alleles [ 44 – 45 ].…”

“…SSR markers can be used for ploidy identification provided that they are variable enough to reveal as many alleles as expected for the ploidy level of the species in some loci. Thus we further evaluated the polymorphism information content ( PIC ) value [ 40 ] using the formula described in Botstein et al [ 41 ]. Following these steps, five EST-SSR primers (MATS2-33, MATS2-48, MATS2-55, MATS2-65 and MATS2-74) that amplified distinct alleles that can be easily recorded with higher PIC values were selected.…”