# Genotyping-by-Sequencing Reveals Marker-Based Genome Stability in Tetraploid Clementines for Triploid Breeding

## TL;DR

This study uses genotyping-by-sequencing to assess genome stability in tetraploid clementines, finding that three tetraploidization methods preserve chromosome structure, making them suitable for triploid breeding.

## Contribution

The study provides genome-wide evidence that three tetraploidization methods maintain genomic stability in citrus, supporting their use in triploid breeding.

## Key findings

- No major genomic gains or losses were detected in tetraploid clementine plants using GBS.
- Three tetraploidization methods preserved chromosome structure at the resolution of GBS markers.
- GBS-based analysis confirmed the suitability of these methods for citrus triploid breeding.

## Abstract

Tetraploid non-apomictic citrus genotypes are key female parents for 4x × 2x hybridizations aimed at producing seedless triploid hybrids. However, the extent to which different tetraploidization methods affect genome integrity remains insufficiently characterized at a genome-wide scale. In this study, genotyping-by-sequencing (GBS) was used to evaluate marker-based genomic stability in ten tetraploid plants of ‘Clemenules’, ‘Fina’, and ‘Marisol’ clementines obtained via colchicine treatment, in vitro adventitious organogenesis, or somatic cybridization. Diploid parental plants, two haploid plants of ‘Clemenules’ and ‘Fina’ clementines, and one doubled haploid plant of ‘Clemenules’ clementine were included, being the haploid and double haploid essential to resolve allelic phases. After quality filtering, 3333 SNP (Single Nucleotide Polymorphism) markers distributed across the nine citrus chromosomes were identified and used to compare allele dosage patterns along the genome. Across all GBS-covered regions, no major marker-based genomic gains or losses were detected in any tetraploid plant. These results indicate that, at the resolution provided by GBS, all three tetraploidization methods largely preserve chromosome structure, supporting their suitability for citrus triploid breeding programs based on 4x × 2x sexual hybridizations.