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Molecular Breeding

, 39:12 | Cite as

Genetic characterization and fine mapping BrCER4 in involved cuticular wax formation in purple cai-tai (Brassica rapa L. var. purpurea)

  • Canjie Wang
  • Honglian Li
  • Yixiao Li
  • Qiufeng Meng
  • Fei Xie
  • Yuejin Xu
  • Zhengjie WanEmail author
Article
  • 38 Downloads

Abstract

Plant aerial organs are covered by waxy cuticles, which are complex mixtures of very long-chain fatty acids and their derivatives. This waxy cuticle can reduce uncontrolled water loss and environmental damage, but a lack of waxy cuticle generates a glossy phenotype, a favorable trait for many crops. The mechanism underlying the glossy phenotype of purple cai-tai (Brassica rapa L. var. purpurea), a unique stem vegetable, has not yet been studied. Genetic analysis indicated that the waxy cuticle in purple cai-tai is controlled by a single locus located on chromosome A01. The trait was ultimately mapped between two markers that are approximately 310 kb apart. Seventy-five genes were identified in the target region. Based on gene annotation, two genes in this region, Bra011487 and Bra011470, are known to be involved in the cuticular wax biosynthesis pathway. No sequence variation for the Bra011487 gene was found between the two parents, while Bra011470 had a 39 bp deletion in the glossy parent. Bra011470 (BrCER4) is a homolog of CER4 (ECERIFERUM4) in Arabidopsis. It was expressed in the leaves, stems, flowers and siliques, and its expression was decreased in the glossy mutant. Analysis with transmission electron microscopy revealed that the cuticular ultrastructure of the mutant leaves and stem was dramatically altered compared with that of the wild-type leaves and stem. Analysis of the cuticular wax of mutant stems further confirmed a severe decrease in primary alcohols. These results indicated that BrCER4 is responsible for surface wax biosynthesis and that a loss-of-function mutation in this gene results in the glossy phenotype. A genetic marker flanking the potentially causal deletion was developed and may be useful for marker-assisted selection in future breeding programs for B. rapa.

Keywords

Purple cai-tai Glossy mutant Cuticular wax BrCER4 

Notes

Funding information

This research was supported by the National Key Research and Development Program of China (2017YFD010803), the China Agriculture Research System (CARS-24-A-06).

Supplementary material

11032_2018_919_MOESM1_ESM.doc (28 kb)
Online Resource 1 Primers used in the present study (DOC 28 kb)
11032_2018_919_MOESM2_ESM.doc (34 kb)
Online Resource 2 The natural cultivars used in the present study (DOC 33 kb)
11032_2018_919_Fig5_ESM.png (232 kb)
Online Resource 3

Permeability of the glossy mutant and WT cuticles. The experiment was repeated twice with similar results. a. Stem water loss rates recorded over 360 min, measured as a percentage of the initial weight of fully hydrated stems. The values are presented as the means of three replicated assays. Error bars = SD. b. Chlorophyll leaching rates recorded over 120 min, measured as a percentage of the total extracted chlorophyll. (PNG 231 kb)

11032_2018_919_MOESM3_ESM.tif (42 kb)
High resolution image (TIF 42 kb)
11032_2018_919_Fig6_ESM.png (668 kb)
Online Resource 4

The DNA sequence information of BrCER4 in wild type and glossy type in purple cai-tai. The gray background represent exons. (PNG 668 kb)

11032_2018_919_MOESM4_ESM.tif (133 kb)
High resolution image (TIF 133 kb)
11032_2018_919_MOESM5_ESM.doc (36 kb)
Online Resource 5 The 39 bp deletion causes a premature stop codon in four transcripts due to a frameshift. BrCER4 represents the constitutive splicing pattern; Brcer4-D1, Brcer4-D2, Brcer4-D3, Brcer4-D4 indicate alternative splicing patterns. The solid boxes represent exons; the gray boxes represent exons that are missing completely or partly compared with those of the constitutively spliced transcripts; the dotted lines denote introns that are missing completely or partly compared with the those of the constitutively spliced transcripts; the continuous lines indicate retention of intron parts; the numbers 59, 80 and 325 indicate the number of nucleotides retained in intron 3; and the dotted boxes indicate premature stop codons. (DOC 36 kb)
11032_2018_919_Fig7_ESM.png (161 kb)
Online Resource 6

Validation of the gene-specific marker on BrCER4 in an F2 population and ten cultivars of purple cai-tai used for the validation of the BrCER4 gene. 1–5: inbred lines (waxy) H1, H2, H3, H7, H10; 6–10: inbred lines (glossy) H5, H6, H8, H16, H20. (PNG 160 kb)

11032_2018_919_MOESM6_ESM.tif (43 kb)
High resolution image (TIF 42 kb)
11032_2018_919_Fig8_ESM.png (16 kb)
Online Resource 7

Evolutionary relationships of the CER4 homologs in Cruciferae. (PNG 16 kb)

11032_2018_919_MOESM7_ESM.tif (768 kb)
High resolution image (TIF 767 kb)

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Canjie Wang
    • 1
    • 2
  • Honglian Li
    • 1
  • Yixiao Li
    • 1
  • Qiufeng Meng
    • 3
  • Fei Xie
    • 1
  • Yuejin Xu
    • 1
  • Zhengjie Wan
    • 1
    Email author
  1. 1.College of Horticulture and Forestry, Key Laboratory of Horticultural Plant Biology, Ministry of EducationHuazhong Agricultural UniversityWuhanChina
  2. 2.Tianmen Vegetable Management OfficeTianmenChina
  3. 3.Ningbo Academy of Agricultural SciencesNingboChina

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