A rare SNP mutation in Brachytic2 moderately reduces plant height and increases yield potential in maize

Abstract

Plant height has long been an important agronomic trait in maize breeding. Many plant height QTLs have been reported, but few of these have been cloned. In this study, a major plant height QTL, qph1, was mapped to a 1.6kb interval in Brachytic2 (Br2) coding sequence on maize chromosome 1. A naturally occurring rare SNP in qph1, which resulted in an amino acid substitution, was validated as the causative mutation. QPH1 protein is located in the plasma membrane and polar auxin transport is impaired in the short near-isogenic line RIL88(qph1). Allelism testing showed that the SNP variant in qph1 reduces longitudinal cell number and decreases plant height by 20% in RIL88(qph1) compared to RIL88(QPH1), and is milder than known br2 mutant alleles. The effect of qph1 on plant height is significant and has no or a slight influence on yield in four F2 backgrounds and in six pairs of single-cross hybrids. Moreover, qph1 could reduce plant height when heterozygous, allowing it to be easily employed in maize breeding. Thus, a less-severe allele of a known dwarf mutant explains part of the quantitative variation for plant height and has great potential in maize improvement.

Keywords: Brachytic2; maize (Zea mays); major QTL; mild mutation; plant height; rare allele..

Fig. 1.

Plant height and ear height variation in BC₄F_2:3. Blue, green and red bars represent the number of qph1/qph1, QPH1/qph1, and QPH1/QPH1 individuals in the BC₄F₃ population developed from RIL88 and RIL279. Plant height (A) and ear height (B) segregations are consistent with the segregation of a single Mendelian factor.

Fig. 2.

Plant height and internode length of RIL88_(QPH1) (BC₆F₃) and RIL88_(qph1). (A) RIL88_(qph1) (two rows on the left), RIL88_(QPH1) (two rows on the right), and their F₁ hybrid (two rows in the middle). Plant height of the hybrid is between the two parental lines and slightly shorter than RIL88_(QPH1). (B) Plant height and ear height comparison between RIL88_(QPH1) (left) and RIL88_(qph1) (right). (C) Internode length comparison between RIL88_(QPH1) (left) and RIL88_(qph1) (right); a greater decrease in internode length from the top of the plant down is observed between near-isogenic lines.

Fig. 3.

Scanning electron microscopy examination of the second internodes from RIL88_(qph1) and RIL88_(QPH1). The second internodes of near-isogenic lines in mid-elongation stage were subjected to histological analysis. (A, B) Longitudinal view of the parenchyma cells of RIL88_(QPH1) and RIL88_(QPH1). (C, D) Transverse view of the parenchyma cells of RIL88_(qph1) and RIL88_(QPH1). (E, F) Transverse view of the epidermal region of RIL88_(qph1) and RIL88_(QPH1).

Fig. 4.

Map-based cloning of qph1. (A) Locations of markers used in BC₄F₂ and BC₅F₂ populations. Number of recombinants and crossover rates between markers are shown before and after the comma. (B) Recombinants identified in BC₄F₂ and BC₅F₂. White, black, and grey indicate homozygous RIL88_(qph1), homozygous RIL88_(QPH1), and heterozygous genotypes. Plant height of recombinant individuals and their progenies in two locations are shown. T, tall; S, short. (C) Fine mapping of qph1 using the BC₆F₂ population. MHC412 and umc2396 were used as flanking markers to identify recombinants. Recombinants were sequenced to determine crossover break points, allowing the qph1 interval to be narrowed down to 4.9kb between SNPs 5968 and 4342. (D) Sequence analysis of the qph1 target region. The qph1 interval locates in exon 5 of the Br2 gene. Five SNPs were identified within it; only SNP5259 caused an amino acid substitution from arginine to leucine. Nucleotides and amino acids in RIL88_(qph1) and RIL88_(QPH1) for polymorphic loci are shown before and after the slash.

Fig. 5.

Validation of qph1 and its functional site through transformation of Arabidopsis homologous mutant atpap1-2. (A) Plant height of the atpgp1-2 mutant transformed with pBI121 empty vector, qph1, mqph1 (mutated qph1), and QPH1. (B) Coleoptile length of the atpgp1-2 mutant transformed with pBI121 empty vector, qph1, mqph1 (mutated qph1), and QPH1. (C) Plant height of qph1, mqph1 (mutated qph1), and QPH1 transformation plants. **, significant difference (0.01 level). Data is expressed as mean ± SD.

Fig. 6.

Transport of [³H] IAA in coleoptiles of RIL88_(QPH1), RIL88_(qph1), and the br2 mutant 114F. CPM, counts per minute. Error bars indicate SD, n=8.

Fig. 7.

Plant height analysis of the four F₂ populations and six pairs of single-cross hybrids. (A) Blue, red, and green represent the plant height of the qph1/qph1, QPH1/qph1, and QPH1/QPH1 individuals, respectively. Phenotype data were collected from two replicates in Hainan (2009, left) and Beijing (2010, right). 50, 100, and 50 individuals of qph1/qph1, QPH1/qph1, and QPH1/QPH1 were planted and analysed, respectively, for each population. (B) Plant height of RIL88_(qph1) (blue) and RIL88_(QPH1) (red) derived single-cross hybrids; 30–40 individuals were planted and analysed, respectively, in each genotype class, and data was collected from two replicates for each hybrid. **, significant difference (0.01 level). Data are expressed as mean ± SD.

Fig. 8.

Protein structure simulation of qph1 generated using Pymol. The six active sites of the protein are shown in yellow; the Arginine to Leucine amino acid substitution on the ninth α-helix in the transmembrane domain is indicated in blue. TMs, transmembrane domains; NBDs, nucleotide binding domains.