Genetic Mapping of Chartreuse Bulb Color in Onion

in Journal of the American Society for Horticultural Science

The most common bulb colors of onion (Allium cepa) are red, yellow, and white; chartreuse is a relatively rare bulb color conditioned by the homozygous recessive genotype at the G locus. In this research, plants with chartreuse bulbs were crossed with inbreds with yellow bulbs to develop segregating families for genetic mapping of the G locus. For all of 17 F2 families, segregations for yellow vs. chartreuse bulbs fit the expected 3:1 ratio (P > 0.05). DNAs were isolated from one F2 family and genotyped for single nucleotide polymorphisms (SNPs) to produce a genetic map of the G locus and 380 SNPs, of which 119 SNPs have not been previously mapped. Segregations for yellow vs. chartreuse bulbs placed the G locus at the end of chromosome 7 at 6.7 cM from the nearest SNP (isotig28625_2789). This codominant SNP marker linked to the G locus should be useful for introgression of recessive chartreuse bulb color into diverse onion populations for commercial production of this uniquely colored onion.

Abstract

The most common bulb colors of onion (Allium cepa) are red, yellow, and white; chartreuse is a relatively rare bulb color conditioned by the homozygous recessive genotype at the G locus. In this research, plants with chartreuse bulbs were crossed with inbreds with yellow bulbs to develop segregating families for genetic mapping of the G locus. For all of 17 F2 families, segregations for yellow vs. chartreuse bulbs fit the expected 3:1 ratio (P > 0.05). DNAs were isolated from one F2 family and genotyped for single nucleotide polymorphisms (SNPs) to produce a genetic map of the G locus and 380 SNPs, of which 119 SNPs have not been previously mapped. Segregations for yellow vs. chartreuse bulbs placed the G locus at the end of chromosome 7 at 6.7 cM from the nearest SNP (isotig28625_2789). This codominant SNP marker linked to the G locus should be useful for introgression of recessive chartreuse bulb color into diverse onion populations for commercial production of this uniquely colored onion.

The color of onion (Allium cepa) bulbs is one of the most important consumer traits, and bulb colors can be white, yellow, red, or chartreuse (El-Shafie and Davis, 1967). White bulbs are conditioned by a dominant allele at the inhibitor (I) locus or the homozygous recessive genotype at the color (C) locus, regardless of genotypes at other color loci. Red bulbs are conditioned by dominant alleles at all of the C, R, and L loci (El-Shafie and Davis, 1967). Khar et al. (2008) and Duangjit et al. (2014) described a second locus (L2) linked to L at which a dominant allele interacts with the R locus to condition red bulbs. Yellow bulbs have a dominant allele at C and the homozygous recessive genotype at one of the loci conditioning red bulbs. The G locus conditions “golden yellow” bulbs when the plant is iiC- (El-Shafie and Davis, 1967), although it is not clear how this color differs from yellow. The homozygous recessive genotype at G conditions chartreuse (light green) bulb color (Fig. 1) in plants that are iiC- regardless of genotypes at the L and R loci (El-Shafie and Davis, 1967). Chartreuse bulbs have been reported in the onion cultivars Australian Brown, Greenella, and Giza #6 (Green et al., 1997; Jones and Mann, 1963). In this research, families segregating for chartreuse bulb color were developed and the G locus mapped relative to SNPs. Molecular markers linked to the G locus should be useful for introgression of recessively inherited chartreuse bulb color into diverse onion populations.

Fig. 1.
Fig. 1.

Chartreuse (left) vs. yellow (right four) bulb colors in onion inbred MSU5718.

Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 145, 2; 10.21273/JASHS04861-20

Methods

Two plants with chartreuse bulbs were identified among progenies from the cross MSU5718A × MSU8155B, which is the female parent of the hybrid ‘Sweet Sandwich’ (Goldman et al., 2001). These two chartreuse plants were intercrossed [breeding plot (BP) 23873]. The cytoplasm and genotype at the nuclear male-fertility restoration (Ms) locus (Jones and Clarke, 1943) of chartreuse plants from BP 23873 were established using pooled DNA (described subsequently) and polymorphisms in the chloroplast accD gene (von Kohn et al., 2013) and nuclear AcPms1 marker (Havey and von Kohn, 2017; Kim et al., 2015), respectively. Single progenies from BP 23873 with chartreuse bulbs and waxy foliage were crossed with inbred B5351 with semiglossy foliage or inbred B9885 with glossy foliage (Damon et al., 2014). Seed was harvested separately from both parents and hybrids identified by yellow bulb color from the chartreuse parent or waxy foliage from B5351 and B9885 parents. Individual hybrid progenies were self-pollinated to produce F2 families. Bulb colors of F2 progenies were scored in field plots as yellow or chartreuse and goodness-of-fit to the expected 3:1 ratio was tested using chi-square analyses. Plants from one F2 family (BP 24334) were self-pollinated and F3 families were grown in field plots and scored for segregations of yellow vs. chartreuse bulb colors. On the basis of segregations in the F3 families, the goodness-of-fit to the expected 1:2:1 ratio for the original F2 family was tested using chi-square analyses.

DNA was isolated from F2 progenies of family BP 24334 using a DNA purification kit (NucleoSpin Plant II Midi; Macherey-Nagel, Düren, Germany) and concentrations established spectrophotometrically (NanoDrop; Thermo Fisher Scientific, Waltham, MA). Single nucleotide polymorphisms (SNPs) were genotyped using DNAs from F2 plants and the Illumina array described by Havey and Ghavami (2018). Goodness-of-fit to the expected 1:2:1 ratio for SNPs were assessed with chi-square analyses and linkages detected at a minimum linkage of logarithm of odds of 12 using Joinmap version 4 (van Ooijen, 2006). Linkage groups were assigned to chromosomes based on 160 SNPs that segregated in earlier mapping populations (Damon and Havey, 2014; Duangjit et al., 2013; Munaiz and Havey, 2020).

Results and Discussion

All of 85 progenies from the initial cross of two chartreuse plants had chartreuse bulbs, as expected because this bulb color is conditioned by the homozygous recessive genotype at the G locus (El-Shafie and Davis, 1967). Polymorphisms in chloroplast accD (von Kohn et al., 2013) and nuclear AcPms1 marker (Havey and von Kohn, 2017; Kim et al., 2015) established that the chartreuse plants selected in this study were S cytoplasmic and possessed dominant alleles at the Ms locus, indicating that the original chartreuse plants likely originated from self-pollination within cytoplasmic male-sterile line MSU5718A due to the presence of male-fertility restoration allele(s) at the Ms (Jones and Clarke, 1943).

Segregations of yellow vs. chartreuse bulbs fit the expected 3:1 ratio for all of 17 F2 families from crosses of chartreuse plants with inbreds B5351 and B9885 (Table 1), in agreement with the genetic model proposed by El-Shafie and Davis (1967). For F3 families from the cross of a chartreuse plant with B5351, 22 families had only chartreuse bulbs, 46 segregated for chartreuse and yellow bulbs, and 15 families had only yellow bulbs which fit the expected 1:2:1 segregation [P = 0.340 (Table 2)]. A total of 1624 SNPs (Havey and Ghavami, 2018) were genotyped using DNAs from 92 F2 progenies, of which 380 segregated and all fit the expected 1:2:1 ratio at P > 0.01 (Table 2). The map locations of 119 previously unmapped SNPs were established (indicated with bold text in Table 2). It was surprising that segregations for bulb colors (Table 1) and all SNPs (Table 2) fit the expected segregation ratios, which may have resulted in parents well adapted to growing conditions in Wisconsin and the robustness of SNP genotyping using the Illumina platform and DNAs from individual plants.

Table 1.

Observed segregations and probabilities of goodness-of-fit to expected 3:1 ratio for yellow vs. chartreuse (Char) bulbs in F2 families of onion.

Table 1.
Table 2.

Chromosome (Chrom), position in centiMorgans, observed (Obs) segregations, and probabilities of goodness-of-fit to expected 1:2:1 ratios for single nucleotide polymorphisms (SNPs) and chartreuse vs. yellow bulb colors in an F2 family of onion.

Table 2.

The G locus mapped to the end of chromosome 7 at 6.7 cM distal from SNP isotig28625_2789 (Table 2). Other major loci conditioning bulb colors in onion have been mapped; the C locus maps to chromosome 6, L and L2 to chromosome 4, and G and R to chromosome 7 (Khar et al., 2008; Masuzaki et al., 2006) (Table 2). The I locus showed linkage at 21 cM with ACM006 (Khar et al., 2008), a simple sequence repeat which has not been placed on the genetic map of onion. Khar et al. (2008) reported that the candidate gene for the R locus [dihydroflavonol 4-reductase (Kim et al., 2004)] mapped 11.1 cM from a restriction fragment length polymorphism revealed by clone AOB212 (King et al., 1998) on chromosome 7. In the genetic map developed by Duangjit et al. (2013), AOB212 mapped 11.2 cM from SNP isotig37252_318 and 27.4 cM from isotig25801_1760. Both of these SNPs segregated in the chartreuse × B5351 map (Table 2), and comparison of these marker positions and orientations indicate that the G locus should map ≈12 cM from R on chromosome 7.

The colors of vegetables impact consumer acceptance, and consumers may respond positively to uniquely colored vegetables as long as other sensorial characteristics (e.g., flavor, texture, aroma) are acceptable (Leksrisompong et al., 2012). Chartreuse onions rarely appear in markets and may be attractive to consumers interested in uniquely colored vegetables. The SNP marker linked with the G locus on chromosome 7 should be useful for backcrossing recessively inherited chartreuse bulb color into elite onion populations, avoiding the need to self-pollinate after each backcross generation to identify heterozygotes and expediting commercial production of onions with this unique bulb color.

Literature Cited

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    • Search Google Scholar
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  • DuangjitJ.BohanecB.ChanA.P.TownC.T.HaveyM.J.2013Transcriptome sequencing to produce SNP-based genetic maps of onionTheor. Appl. Genet.12620932101

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    • Search Google Scholar
    • Export Citation
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    • Search Google Scholar
    • Export Citation
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    • Search Google Scholar
    • Export Citation
  • KingJ.J.BradeenJ.M.BarkO.McCallumJ.A.HaveyM.J.1998A low-density genetic map of onion reveals a role for tandem duplication in the evolution of an extremely large diploid genomeTheor. Appl. Genet.965262

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  • LeksrisompongP.P.WhitsonM.E.TruongV.D.DrakeM.A.2012Sensory attributes and consumer acceptance of sweet potato cultivars with varying flesh colorsJ. Sens. Stud.275969

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  • MasuzakiS.ShigyoM.YamauchiN.2006Complete assignment of structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of shallot (Allium cepa L.)Genes Genet. Syst.81255263

    • Search Google Scholar
    • Export Citation
  • MunaizE.D.HaveyM.J.2020Genetic analyses of epicuticular waxes associated with the glossy foliage of ‘White Persian’ onionJ. Amer. Soc. Hort. Sci.1456772

    • Search Google Scholar
    • Export Citation
  • van OoijenJ.W.2006JoinMap 4 software for the calculation of genetic linkage maps in experimental populations. Kyazma Wageningen the Netherlands

  • von KohnC.KiełkowskaA.HaveyM.J.2013Sequencing and annotation of the chloroplast DNAs of normal (N) male-fertile and male-sterile (S) cytoplasms of onion and single nucleotide polymorphisms distinguishing these cytoplasmsGenome56737742

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Contributor Notes

I thank Christy Stewart and Franco Parisi for technical assistance.Names are necessary to report factually on available data; however, the U.S. Department of Agriculture (USDA) neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable.M.J.H. is the corresponding author. E-mail: michael.havey@usda.gov.
  • View in gallery

    Chartreuse (left) vs. yellow (right four) bulb colors in onion inbred MSU5718.

  • DamonS.HaveyM.J.2014Quantitative trait loci controlling amounts and types of epicuticular waxes in onionJ. Amer. Soc. Hort. Sci.139597602

    • Search Google Scholar
    • Export Citation
  • DamonS.GrovesR.HaveyM.J.2014Variation for epicuticular waxes on onion foliage and impacts on numbers of onion thripsJ. Amer. Soc. Hort. Sci.139495501

    • Search Google Scholar
    • Export Citation
  • DuangjitJ.BohanecB.ChanA.P.TownC.T.HaveyM.J.2013Transcriptome sequencing to produce SNP-based genetic maps of onionTheor. Appl. Genet.12620932101

    • Search Google Scholar
    • Export Citation
  • DuangjitJ.WelshK.WiseM.BohanecB.HaveyM.J.2014Genetic analyses of anthocyanin concentrations and intensity of red-bulb color among segregating haploid progenies of onionMol. Breed.347585

    • Search Google Scholar
    • Export Citation
  • El-ShafieM.DavisG.1967Inheritance of bulb color in Allium cepaHilgardia9607622

  • GreenF.N.BaurR.ThomsonM.McCarthyL.1997An example of chartreuse skin colour in onion (Allium cepa L.) cultivar GreenellaGenet. Resources Crop Evol.44491493

    • Search Google Scholar
    • Export Citation
  • GoldmanI.L.SchroeckG.HaveyM.J.2001History of public onion breeding programs and pedigree of public onion germplasm releases in the United StatesPlant Breed. Rev.2067103

    • Search Google Scholar
    • Export Citation
  • HaveyM.J.von KohnC.2017Efficacy of molecular markers jnurf13 and AcPms1 for prediction of genotypes at the nuclear Ms locus in North American open-pollinated populations of onionHortScience5210521053

    • Search Google Scholar
    • Export Citation
  • HaveyM.J.GhavamiF.2018Informativeness of single nucleotide polymorphisms and relationships among onion populations from important world production regionsJ. Amer. Soc. Hort. Sci.1433444

    • Search Google Scholar
    • Export Citation
  • JonesH.A.ClarkeA.1943Inheritance of male sterility in the onion and the production of hybrid seedProc. Amer. Soc. Hort. Sci.43189194

  • JonesH.A.MannL.K.1963Onions and their allies. Interscience New YorkNY

  • KharA.JakšeJ.HaveyM.J.2008Segregations for onion-bulb colors reveal that red is controlled by at least three lociJ. Amer. Soc. Hort. Sci.1334247

    • Search Google Scholar
    • Export Citation
  • KimS.KimC.-W.ParkM.ChoiD.2015Identification of candidate genes associated with fertility restoration of cytoplasmic male sterility in onion (Allium cepa L.) using a combination of bulked segregant analysis and RNA-seqTheor. Appl. Genet.12822892299

    • Search Google Scholar
    • Export Citation
  • KimS.BinzelM.L.ParkS.YooK.S.PikeL.M.2004Inactivation of DFR (Dihydroflavonol 4-reductase) gene transcription results in blockage of anthocyanin production in yellow onions (Allium cepa)Mol. Breed.14253263

    • Search Google Scholar
    • Export Citation
  • KingJ.J.BradeenJ.M.BarkO.McCallumJ.A.HaveyM.J.1998A low-density genetic map of onion reveals a role for tandem duplication in the evolution of an extremely large diploid genomeTheor. Appl. Genet.965262

    • Search Google Scholar
    • Export Citation
  • LeksrisompongP.P.WhitsonM.E.TruongV.D.DrakeM.A.2012Sensory attributes and consumer acceptance of sweet potato cultivars with varying flesh colorsJ. Sens. Stud.275969

    • Search Google Scholar
    • Export Citation
  • MasuzakiS.ShigyoM.YamauchiN.2006Complete assignment of structural genes involved in flavonoid biosynthesis influencing bulb color to individual chromosomes of shallot (Allium cepa L.)Genes Genet. Syst.81255263

    • Search Google Scholar
    • Export Citation
  • MunaizE.D.HaveyM.J.2020Genetic analyses of epicuticular waxes associated with the glossy foliage of ‘White Persian’ onionJ. Amer. Soc. Hort. Sci.1456772

    • Search Google Scholar
    • Export Citation
  • van OoijenJ.W.2006JoinMap 4 software for the calculation of genetic linkage maps in experimental populations. Kyazma Wageningen the Netherlands

  • von KohnC.KiełkowskaA.HaveyM.J.2013Sequencing and annotation of the chloroplast DNAs of normal (N) male-fertile and male-sterile (S) cytoplasms of onion and single nucleotide polymorphisms distinguishing these cytoplasmsGenome56737742

    • Search Google Scholar
    • Export Citation
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