Abstract
Most indigenous citrus varieties are assumed to be natural hybrids, but their parentage has so far been determined in only a few cases because of their wide genetic diversity and the low transferability of DNA markers. Here we infer the parentage of indigenous citrus varieties using simple sequence repeat and indel markers developed from various citrus genome sequence resources. Parentage tests with 122 known hybrids using the selected DNA markers certify their transferability among those hybrids. Identity tests confirm that most variant strains are selected mutants, but we find four types of kunenbo (Citrus nobilis) and three types of tachibana (Citrus tachibana) for which we suggest different origins. Structure analysis with DNA markers that are in Hardy–Weinberg equilibrium deduce three basic taxa coinciding with the current understanding of citrus ancestors. Genotyping analysis of 101 indigenous citrus varieties with 123 selected DNA markers infers the parentages of 22 indigenous citrus varieties including Satsuma, Temple, and iyo, and single parents of 45 indigenous citrus varieties, including kunenbo, C. ichangensis, and Ichang lemon by allele-sharing and parentage tests. Genotyping analysis of chloroplast and mitochondrial genomes using 11 DNA markers classifies their cytoplasmic genotypes into 18 categories and deduces the combination of seed and pollen parents. Likelihood ratio analysis verifies the inferred parentages with significant scores. The reconstructed genealogy identifies 12 types of varieties consisting of Kishu, kunenbo, yuzu, koji, sour orange, dancy, kobeni mikan, sweet orange, tachibana, Cleopatra, willowleaf mandarin, and pummelo, which have played pivotal roles in the occurrence of these indigenous varieties. The inferred parentage of the indigenous varieties confirms their hybrid origins, as found by recent studies.
Highlights
The genus Citrus L. (Family Rutaceae, subfamily Aurantiodeae) covers a wide range of edible and commercial varieties, including sweet orange, lemon, lime, grapefruit, and mandarins such as Clementine, Satsuma, King, and ponkan [1,2,3,4]
Another clustering analysis of whole genome shotgun sequences of sweet orange ‘Ridge Pineapple’ yielded 381,909 consensus sequences from 866,700 reads, but 46,341 Clementine BAC end sequences were not used for assembly because of their low redundancy
simple sequence repeat (SSR) mining of the Clementine haploid genome sequence [43] identified 310,413 candidate SSR regions for both v0.9 and v1.0 genomes. These candidate regions were verified with resequencing data obtained from Next generation sequencing (NGS) analysis of 15 citrus varieties (banpeiyu A004, Clementine A009, dancy A016, hyuganatsu A036 and A038, King A054, Kishu A066, ponkan A108, Satsuma A113 and A122, sweet orange A162, willowleaf (Mediterranean) mandarin A200, ‘Encore’ B014, ‘Harehime’ B017, and ‘Kiyomi’ tangor B031)
Summary
The genus Citrus L. (Family Rutaceae, subfamily Aurantiodeae) covers a wide range of edible and commercial varieties, including sweet orange, lemon, lime, grapefruit, and mandarins such as Clementine, Satsuma, King, and ponkan [1,2,3,4]. The production of major citrus varieties in tropical to sub-tropical and temperate zones exceeds 90 million tons, and the citrus industry occupies a significant position in the fruit industry and in global agriculture [5,6]. Wide genetic diversity observed in Citrus, has made it difficult for taxonomists to draw a clear picture of their classification. Mutants have occasionally been selected from limb sports or nucellar seedlings, and these constitute large variant strains [2,8,9,10]. Understanding how these modern citrus varieties arose from the ancestral basic species would bring us important insights for future citrus breeding
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