Characterization and Mapping of a Rolling Leaf Mutant Allele rlT73 on Chromosome 1BL of Wheat

Abstract

Leaf rolling is regarded as an important morphological trait in wheat breeding. Moderate leaf rolling is helpful to keep leaves upright and improve the photosynthesis of plants, leading to increased yield. However, studies on the identification of genomic regions/genes associated with rolling leaf have been reported less frequently in wheat. In this study, a rolling leaf mutant, T73, which has paired spikelets, dwarfism, and delayed heading traits, was obtained from a common wheat landrace through ethyl methanesulfonate mutagenesis. The rlT73 mutation caused an increase in the number of epidermal cells on the abaxial side and the shrinkage of bulliform cells on the adaxial side, leading to an adaxially rolling leaf phenotype. Genetic analysis showed that the rolling leaf phenotype was controlled by a single recessive gene. Further Wheat55K single nucleotide polymorphism array-based bulked segregant analysis and molecular marker mapping delimited rlT73 to a physical interval of 300.29-318.33 Mb on the chromosome arm 1BL in the Chinese Spring genome. We show that a point mutation at the miRNA165/166 binding site of the HD zipper class III transcription factor on 1BL altered its transcriptional level, which may be responsible for the rolling leaf phenotype. Our results suggest the important role of rlT73 in regulating wheat leaf development and the potential of miRNA-based gene regulation for crop trait improvement.

Keywords: HD zipper class III transcription factor; miRNA; rolling leaf; wheat.

Figure 1

Phenotypic characterization of T73 mutant plants. (A) CS and T73 plants grown in a field environment. (B) Spike morphology of CS and T73. (C) Seeds of CS and T73. (D) Statistics of plant height, spike length, spikelet number, grain number per spike, tiller number, flag leaf length, thousand grain weight, grain length, and grain width of CS and T73. The values for grain length and grain width were an average of 50 grains. * p < 0.05; ** p < 0.01.

Figure 2

Phenotype changes of CS and T73 leaves. (A) Plant stature (top) and transverse leaf sections (bottom) of CS and T73. (B) Toluidine blue-stained cross sections of CS and T73 leaves. BC, bulliform cell; VB, vascular bundles; SC, sclerenchyma cells; EC, epidermal cell; Ad, adaxial; and Ab, abaxial.

Figure 3

Genetic characterization of rlT73. (A) Leaves of CS (left), F₁ of T73 × CS (middle), and T73 (right). (B) Number of SNPs distributed on wheat chromosomes in the T73 × AK58 mapping population. (C) The enrichment of SNPs on wheat chromosomes in the T73 × AK58 mapping population.

Figure 4

Molecular mapping and candidate gene analysis of rlT73. (A) Genetic map of rlT73. (B) the molecular markers used to narrow down the genetic interval of rlT73. 45-6, 448-1, 688-4, 401-6, TC50, and TC54 are homozygous lines that displayed recombinant events in the rlT73 region. The actual number (n) of each recombinant event is listed. R, rolling leaf phenotype; WT, wild-type leaf. (C) Gene structure of TraesCS1B03G0515400. (D) Schematic representation of the missense variant involved in the miRNA166 binding site of TraesCS1B03G0515400. (E) Expression levels of TraesCS1B03G0515400 (HB-B2) and its homeologs (HB-A2 and HB-D2) in wheat leaves of CS and T73. Plants were grown in growth chambers at 22 °C with an 18 h light/6 h dark photoperiod. Leaves were collected at the jointing stage. * p < 0.05.