Identification and Molecular Analysis of Putative Self-Incompatibility Ribonuclease Alleles in an Extreme Polyploid Species, Prunus laurocerasus L.

Abstract

Cherry laurel (Prunus laurocerasus L.) is an extreme polyploid (2n = 22x) species of the Rosaceae family where gametophytic self-incompatibility (GSI) prevents inbreeding. This study was carried out to identify the S-ribonuclease alleles (S-RNases) of P. laurocerasus using PCR amplification of the first and second intron region of the S-RNase gene, cloning and sequencing. A total of 23 putative S-RNase alleles (S1–S20, S5m, S13m, and S18m) were sequenced from the second (C2) to the fifth conserved region (C5), and they shared significant homology to other Prunus S-RNases. The length of the sequenced amplicons ranged from 505 to 1,544 bp, and similar sizes prevented the proper discrimination of some alleles based on PCR analysis. We have found three putatively non-functional alleles (S5m, S18m, and S9) coding for truncated proteins. Although firm conclusions cannot be drawn, our data seem to support that heteroallelic pollen cannot induce self-compatibility in this polyploid Prunus species. The identities in the deduced amino acid sequences between the P. laurocerasus and other Prunus S-RNases ranged between 44 and 100%, without a discontinuity gap separating the identity percentages of trans-specific and more distantly related alleles. The phylogenetic position, the identities in nucleotide sequences of the second intron and in deduced amino acid sequences found one or more trans-specific alleles for all but S10, S14, S18, and S20 cherry laurel RNases. The analysis of mutational frequencies in trans-specific allele pairs indicated the region RC4–C5 accepts the most amino acid replacements and hence it may contribute to allele-specificity. Our results form the basis of future studies to confirm the existence and function of the GSI system in this extreme polyploid species and the alleles identified will be also useful for phylogenetic studies of Prunus S-RNases as the number of S-RNase sequences was limited in the Racemose group of Prunus (where P. laurocerasus belongs to).

S-Locus Genotyping in Japanese Plum by High Throughput Sequencing Using a Synthetic S-Loci Reference Sequence

“…The alleles recovered from the ‘Sanyueli’ reference genome showed lower than expected trans-specific similarity within Prunus , whereas the S-RNase and SFB recovered from the ‘ZGL’ reference genome were highly similar to S06 S-RNase from P. speciosa and SFB5 from P. tenella , respectively. High similarity between alleles of different species compared to those of the same species—called trans-specific polymorphism—is a common phenomenon within GSI [ 12 , 38 , 39 , 40 ]. Sequence analysis of both genes showed typical structural arrangements and size ranges as in other P. salicina alleles [ 35 , 41 , 42 , 43 , 44 , 45 , 46 ].…”

“…In Prunus sp., as in other species of the Rosaceae, pollen allele specificity is determined by an F-box protein (SFB) that is expressed in the pollen grain [ 7 ], and pistil allele specificity is determined by a ribonuclease (S-RNase) that is expressed in styles [ 8 , 9 ], and inhibits the growth of self-pollen tubes [ 10 ]. The genotyping of S -locus in both natural and domesticated plant species has confirmed the high allelic diversity and sequence polymorphism [ 11 , 12 ].…”

Synergistic approach of PCR-based fragment length analysis and amplicon deep sequencing reveals rich diversity of S-alleles in sweet cherries from the Caucasian region of origin

“…The improved consensus primer pair flanking the first intron of the S-RNase gene, namely PaConsI-F/R2, was suggested for high throughput S-allele genotyping (Sonneveld et al, 2006). This system has become well-established and has been used extensively for S-allele genotyping of sweet cherry (Vaughan et al, 2008;Ganopoulos et al, 2012;Sharma et al, 2016;Cachi et al, 2017;Marchese et al, 2017;Azizi-Gannouni et al, 2018) and has also be transferred to other Prunus species (Halaśz et al, 2021a, Halaśz et al, 2021bNicolaś-Almansa et al, 2023). It further served as a basis for the further development of the genotyping method, such as the one-tube reaction assay described by Cmejlova et al (2023).…”

Insights into tissue-specific anthocyanin accumulation in Japanese plum (Prunus salicina L.) fruits: A comparative study of three cultivars

“…Prunus is a genus with high biodiversity in the form of trees and shrubs, including the fruits of plum, cherry, peach, nectarine, apricot and almond ( Halász et al, 2021 ). The attractive and high economic value fruits of different species belonging to the genus have been extensively studied in recent years, especially in terms of their effects on human health ( Abanoz & Okcu, 2022 ).…”