Plant Biotechnology Journal最新文献

Increasing dietary fibre (DF) intake is an important target to improve health. An attractive strategy for this is to increase DF in wheat which is derived principally from the endosperm cell wall polysaccharide arabinoxylan (AX). The water-extractable form of this (WE-AX) accounts for most soluble dietary fibre (SDF), which is believed to confer particular health benefits. A region of chromosome 6B in some wheat varieties confers high SDF and here we show that the cause is an allele encoding a peroxidase family protein with a single residue change (PER1-v) associated with high WE-AX, compared to the more common form (PER1). Both wheat lines carrying this natural PER1-v variant and those with an induced knockout mutation of PER1 showed reduced dimerization of endosperm ferulate consistent with a mechanism of decreased cross-linking in the cell wall that increases WE-AX. Transiently expressed PER1_RFP fusion protein driven by the native promoter in wheat endosperm was shown to localise to cell walls, whereas PER1-v_RFP did not. We therefore propose that PER1-v lacks the capacity to dimerise AX ferulate in vivo due to mis-localisation caused by the missense single-nucleotide polymorphism (SNP) in the PER1-v allele, so that the SNP acts as a perfect marker. This marker can be used to identify current wheat varieties with high WE-AX to be used by processors and by breeders to ensure future varieties have high WE-AX to make healthier wheat-based foods.

In plants, genetic manipulations are commonly performed using Agrobacterium tumefaciens, a plant pathogen whose abilities as a 'natural genetic engineer' have been co-opted for biotechnological applications. In the widely used binary vector systems, Agrobacterium already contains a plasmid with most of the genes needed for virulence and is transformed with a second vector carrying the DNA of interest that will be transferred to the plant. Many widely used binary vectors have not been substantially improved during the past few decades, however, and often have legacy features that are not needed for many contemporary applications, such as cumbersome cloning sites that introduce sequence 'scars' between proteins of interest and any fusion protein, including GFP. Here, we introduce the assembly vector (AVEC) plasmids, a series of modular vector backbones that are easily customized using inexpensive DNA assembly cloning methods and that are ideal for testing multiple epitope tags, fluorophores, or non-coding sequences, such as promoters or 5'/3' untranslated regions. We show that pAVEC plasmids match or exceed the performance of commonly used binary vectors for transient and transgenic expression of proteins fused to fluorophores and the proximity labelling tool, TurboID. To illustrate the versatility of the pAVEC system, we demonstrate that replacing an herbicide resistance gene with a silencing suppressor gene increases protein expression in cells by an order of magnitude. We anticipate that the pAVEC system will accelerate investigations of plant molecular biology and that the modular, intentional design of these plasmids will be useful to investigate future improvements to binary plasmid design.

Fruit shape is an important external quality trait that directly determines the market value. Modification of fruit shape has emerged as a key focus in crop improvement, but the regulatory network of fruit shape specifications remains largely unknown. Here, we identified a short fruit mutant (sf5) that was caused by a C-to-T single nucleotide polymorphism (SNP) in TONNEAU2 (CsTON2), a microtubule-associated gene encoding the B subunit of protein phosphatase 2A (PP2A). Overexpression of CsTON2 in the sf5 background partially rescued the mutant phenotype, while knockout of CsTON2 led to severe developmental defects and dwarfism. We further demonstrated that CsTON2 physically interacts with CsTRM5 and CsSUN, two key regulators of fruit shape in cucumber. The SNP change of CsTON2 in sf5 mutant impairs the interaction with CsTRM5 and CsSUN, and decreases the protein stability of CsSUN. Genetic analyses revealed that CsTON2, CsTRM5 and CsSUN coordinately regulate fruit shape development by modulating cell division direction in cucumber. Therefore, our findings shed insights into the role of microtubule-associated protein complex in fruit shape determination and provide new gene targets for breeding cucumber varieties with favourable fruit shapes.