Theoretical and Applied Genetics

, Volume 131, Issue 3, pp 673–684 | Cite as

Identification and fine mapping of a stay-green gene (Brnye1) in pakchoi (Brassica campestris L. ssp. chinensis)

  • Nan Wang
  • Zhiyong Liu
  • Yun Zhang
  • Chengyu Li
  • Hui Feng
Original Article

Abstract

Key message

Using bulked segregant analysis combined with next-generation sequencing, we delimited the Brnye1 gene responsible for the stay-green trait of nye in pakchoi. Sequence analysis identified Bra019346 as the candidate gene.

Abstract

“Stay-green” refers to a plant trait whereby leaves remain green during senescence. This trait is useful in the cultivation of pakchoi (Brassica campestris L. ssp. chinensis), which is marketed as a green leaf product. This study aimed to identify the gene responsible for the stay-green trait in pakchoi. We identified a stay-green mutant in pakchoi, which we termed “nye”. Genetic analysis revealed that the stay-green trait is controlled by a single recessive gene, Brnye1. Using the BSA-seq method, a 3.0-Mb candidate region was mapped on chromosome A03, which helped us localize Brnye1 to an 81.01-kb interval between SSR markers SSRWN27 and SSRWN30 via linkage analysis in an F2 population. We identified 12 genes in this region, 11 of which were annotated based on the Brassica rapa annotation database, and one was a functionally unknown gene. An orthologous gene of the Arabidopsis gene AtNYE1, Bra019346, was identified as the potential candidate for Brnye1. Sequence analysis revealed a 40-bp insertion in the second exon of Bra019346 in nye, which generated the TAA stop codon. A candidate gene-specific Indel marker in 1561 F2 individuals showed perfect cosegregation with Brnye1 in the nye mutant. These results provide a foundation for uncovering the molecular mechanism of the stay-green trait in pakchoi.

Notes

Acknowledgements

This work was supported by Grants from the National Natural Science Foundation of China (no. 31601749). We would like to thank Editage for English language editing. We also would like to thank Oebiotech for whole-genome resequencing.

Compliance with ethical standards

Conflict of interest

The authors declare that there are no conflicts of interest in the reported research.

Ethical standards

The authors note that this research was performed and reported in accordance with ethical standards of the scientific conduct.

Supplementary material

122_2017_3028_MOESM1_ESM.tif (2.3 mb)
Supplementary Fig. S1 Gel image of confirmation polymorphism of tightly linked SSR markers between nye and ‘FT’. Note: P1: ‘FT’; P2: nye. (TIFF 2304 kb)
122_2017_3028_MOESM2_ESM.tif (14.5 mb)
Supplementary Fig. S2 Gel images of markers tightly linked to Brnye1. a: Recombinant individuals with SSRWN26 in the mapping population; b: Recombinant individuals with SSRWN25 in the mapping population; c: Recombinant individuals with SSRWN30 in the mapping population; d: Recombinant individuals with SSRWN27 in the mapping population. Note: M: Marker; P1: ‘FT’; P2: nye; *: recombinant individuals. (TIFF 14851 kb)
122_2017_3028_MOESM3_ESM.pdf (21 kb)
Supplementary Fig. S3 Putative ORF of the unknown gene Bra019336 in the 81.01-kb region identified by Softberry. (PDF 21 kb)
122_2017_3028_MOESM4_ESM.tif (9.9 mb)
Supplementary Fig. S4 Sequence alignment of Bra019336 in nye and ‘13A510’. (TIFF 10160 kb)
122_2017_3028_MOESM5_ESM.tif (24.2 mb)
Supplementary Fig. S5 Gel images of polymorphisms of the full length (a), RT-PCR (b), and 3′RACE of Bra019346 (c) between ‘13A510’ and nye; and gel images of the full length of Bra019336 (d) and the promoter of Bra019346 (e) between ‘13A510’ and nye. (TIFF 24768 kb)
122_2017_3028_MOESM6_ESM.tif (20.7 mb)
Supplementary Fig. S6 Sequence alignment of the CDS (a) in ‘13A510’ and nye (TIFF 21176 kb)
122_2017_3028_MOESM7_ESM.tif (4.4 mb)
Supplementary Fig. S6 Partial 3′RACE of Bra019346 (b) in ‘13A510’ and nye (TIFF 4511 kb)
122_2017_3028_MOESM8_ESM.tif (1.4 mb)
Supplementary Fig. S7 Gel images of the indel marker of the bands cosegregated with the mutant nye. Note: M: Marker; P1: ‘FT’; P2: nye. (TIFF 1406 kb)
122_2017_3028_MOESM9_ESM.tif (52.4 mb)
Supplementary Fig. S8 Sequence alignment of the promoter of Bra019346 in ‘13A510’ and nye. (TIFF 53618 kb)
122_2017_3028_MOESM10_ESM.tif (603 kb)
Supplementary Fig. S9 Protein conservative domains analysis of Bra019346. (TIFF 602 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2017

Authors and Affiliations

  1. 1.Liaoning Key Laboratory of Genetics and Breeding for Cruciferous Vegetable CropsShenyang Agricultural UniversityShenyangChina
  2. 2.Department of HorticultureShenyang Agricultural UniversityShenyangChina

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