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Identification of a candidate gene responsible for the G locus determining chartreuse bulb color in onion (Allium cepa L.) using bulked segregant RNA-Seq

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Abstract

Key message

A gene encoding a laccase responsible for chartreuse onion bulb color was identified. Markers tagging this gene showed perfect linkage with bulb colors among diverse germplasm.

Abstract

To identify a casual gene for the G locus determining chartreuse bulb color in onion (Allium cepa L.), bulked segregant RNA-Seq (BSR-Seq) was performed using yellow and chartreuse individuals of a segregating population. Through single nucleotide polymorphism (SNP) and differentially expressed gene (DEG) screening processes, 163 and 143 transcripts were selected, respectively. One transcript encoding a laccase-like protein was commonly identified from SNP and DEG screening. This transcript contained four highly conserved copper-binding domains known to be signature sequences of laccases. This gene was designated AcLAC12 since it showed high homology with Arabidopsis AtLAC12. A 4-bp deletion creating a premature stop codon was identified in exon 5 of the chartreuse allele. Another mutant allele in which an intact LTR-retrotransposon was transposed in exon 5 was identified from other chartreuse breeding lines. Genotypes of molecular markers tagging AcLAC12 were perfectly matched with bulb color phenotypes in segregating populations and diverse breeding lines. All chartreuse breeding lines contained inactive alleles of DFR-A gene determining red bulb color, indicating that chartreuse color appeared when both DFR-A and AcLAC12 genes were inactivated. Linkage maps showed that AcLAC12 was positioned at the end of chromosome 7. Transcription levels of structural genes encoding enzymes in anthocyanin biosynthesis pathway were generally reduced in chartreuse bulk compared with yellow bulk. Concentrations of total quercetins were also reduced in chartreuse onion. However, significant amounts of quercetins were detected in chartreuse onion, implying that AcLAC12 might be involved in modification of quercetin derivatives in onion.

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Data availability

Nucleotide sequences of AcLAC12 alleles are accessible at NCBI Database under the accession numbers from OK33673 to OK336705.

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Acknowledgements

This work was supported by “Cooperative research program for agriculture science and technology development (Project No. PJ0161682021)” funded by Rural Development Administration, Republic of Korea. It was also supported by Golden Seed Project (Center for Horticultural Seed Development, No 213007-05-5-SBB10) of Korea Institute of Planning and Evaluation for Technology in Food, Agriculture and Forestry (IPET). The authors thank Nari Yu, Ji-hwa Heo, Jeong-An Yoo, and Su-jeong Kim for their dedicated technical assistance.

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Authors and Affiliations

  1. Department of Horticulture, Biotechnology Research Institute, Chonnam National University, Gwangju, 61186, Republic of Korea

    SeongChan Jeon & Sunggil Kim

  2. National Institute of Horticultural and Herbal Science, RDA, Muan, 58545, Republic of Korea

    JiWon Han & Cheol-Woo Kim

  3. Department of Food Science and Technology, Chonnam National University, Gwangju, 61186, Republic of Korea

    Ju-Gyeong Kim & Jae-Hak Moon

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  1. SeongChan Jeon
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Contributions

SeongChan Jeon performed experiments and drafted the manuscript. JiWon Han and Cheol-Woo Kim produced segregating populations. Ju-Gyeong Kim and Jae-Hak Moon performed measurements of concentrations of quercetin derivatives. Sunggil Kim organized and coordinated this research project and edited the final manuscript.

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Correspondence to Sunggil Kim.

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Supplementary Information

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122_2021_4016_MOESM1_ESM.tif

Supplementary Fig. 1 Alignment of trimmed reads produced from chartreuse and yellow bulked RNAs to sequences of AcLAC12. Images of read alignments were produced using IGV software (TIF 122 kb)

122_2021_4016_MOESM2_ESM.tif

Supplementary Fig. 2 Correlations of expression levels of all transcripts between chartreuse and yellow bulked RNAs (TIF 32 kb)

122_2021_4016_MOESM3_ESM.tif

Supplementary Fig. 3 Comparison of expression levels of structural genes encoding enzymes in the flavonoid biosynthesis pathway between chartreuse and yellow bulked RNAs. Transcription levels of genes were estimated by RNA-Seq (TIF 72 kb)

122_2021_4016_MOESM4_ESM.tif

Supplementary Fig. 4 Concentrations of quercetin derivatives in yellow and chartreuse onions. A HPLC peak patterns of yellow, chartreuse, red, and white onions. HPLC analyses of red and white onions performed in a previous study (Seo et al. 2020) were included for comparison with the chartreuse onion. Standard 1: Quercetin 3,7-diglucopyranoside; Standard 2: Quercetin 3,4′-diglucopyranoside; Standard 3: Quercetin 3-glucopyranoside; Standard 4: Quercetin 4'-glucopyranoside; Standard 5: Isorhamnetin 3-glucopyranoside; and Standard 6: Quercetin aglycon. B Estimated concentrations of four quercetin derivatives in yellow, chartreuse, red, and white onions (TIF 205 kb)

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Jeon, S., Han, J., Kim, CW. et al. Identification of a candidate gene responsible for the G locus determining chartreuse bulb color in onion (Allium cepa L.) using bulked segregant RNA-Seq. Theor Appl Genet 135, 1025–1036 (2022). https://doi.org/10.1007/s00122-021-04016-5

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