Biology subjects

Milner, M. J.

Publications and source records attributed to Milner, M. J..

4 recordsLinked to original sources

bioRxiv2026

Tracking Transgenes with Color: RUBY as a Visual Marker in CRISPR-Edited Mutant Plants in Two Triticum Species

CRISPR-Cas9 is a powerful tool for precise genome editing in plants, but the presence of foreign DNA, such as T-DNA, raises regulatory concerns and complicates mutant screening and field studies of edited material. Detecting plants with good transgene expression and later removing the T-DNA from edited plants is both time-consuming and costly. To address this, we developed a system that uses the non-destructive RUBY reporter, linked to the CRISPR-Cas9 cassette, and expressed under the ZmUbi1 promoter. To assess the applicability of the system, it was tested on two Triticum species, targeting three genes in either tetraploid or hexaploid wheat. Strong correlations were observed in both T0 and T1 plants between betalain content and Cas9 expression, allowing for the quick identification of plants likely to be edited. Furthermore, the RUBY reporter could be used to select against the transgenic CRISPR-Cas9 cassette in subsequent generations at both the seed and seedling stages, thereby reducing the number of plants that need to be screened to identify edited lines without a T-DNA. This approach, using a nondestructive reporter, enabled rapid distinction between transgene expression in primary transgenics and served as a negative selection in the T1 generation, streamlining selection towards edited and T-DNA-free progeny.

plant biology↗

bioRxiv2025

Genome assembly and population genomic analysis of Aegilops caudata uncover a rich source for disease and insect pest resistance for wheat improvement

Aegilops caudata L. is a rich source of resistance genes to major wheat pathogens and pests, yet its complex genome structure and lack of a reference sequence have hindered genetic analysis, gene discovery, and introgression. Here, we report a high-quality genome sequence assembly of Ae. caudata accession S740-69, integrated with optical mapping to precisely delineate introgressed segments from structurally altered genomic regions in wheat-Ae. caudata translocation lines carrying a novel stem rust resistance gene. Resequencing and phenotyping of 95 diverse accessions, coupled with k-mer-based association mapping, enabled population-level identification of several novel loci for resistance to wheat pests and diseases, such as greenbug and stem rust. Virus-induced gene silencing confirmed Aecau6C01G127270 (SrAect1) as the first functionally characterized stem rust resistance gene from Ae. caudata. This new genomic framework provides a foundation for systematic mining of resistance genes in this species and their efficient introgression into wheat.

genomics↗

bioRxiv2022

OsPSTOL but not TaPSTOL can play a role in nutrient use efficiency and works through conserved pathways in both wheat and rice

There is a large demand to reduce inputs for current crop production, particularly phosphate and nitrogen inputs which are the two most frequently added supplements to agricultural production. Gene characterization is often limited to the native species from which it was identified, but may offer benefits to other species. To understand if OsPSTOL, a gene identified from rice which improves tolerance to low P growth conditions, might improve performance and provide the same benefit in wheat, OsPSTOL was transformed into wheat and expressed from a constitutive promoter. The ability of OsPSTOL to improve nutrient acquisition under low phosphate or low nitrogen was evaluated. Here we show that OsPSTOL works through a conserved pathway in wheat and rice to improve yields under both low phosphate and low nitrogen. This increase is yield is mainly driven by improved uptake from the soil driving increased biomass and ultimately increased seed number, but does not change the concentration of N in the straw or grain. Overexpression of OsPSTOL in wheat modifies N regulated genes to aid in this uptake whereas the putative homolog TaPSTOL does not suggesting that expression of OsPSTOL in wheat can help to improve yields under low input agriculture.

plant biology↗

bioRxiv2021

Over-expression of the brassinosteroid gene TaDWF4 increases wheat productivity under low and sufficient nitrogen through enhanced carbon assimilation

There is a strong pressure to reduce nitrogen (N) fertiliser inputs while maintaining or increasing current cereal crop yields. Brassinosteroids, (BR), are a group of phytohormones essential for plant growth and development, that have been demonstrated to regulate several agronomic traits. DWF4 encodes a cytochrome P450 that catalyses a rate-limiting step in BR synthesis. We show that overexpression of the dominant shoot expressed homoeologue TaDWF4-B in wheat can increase plant productivity by up to 105% under a range of N levels on marginal soils, resulting in increased N use efficiency (NUE). We show that a two to four-fold increase in TaDWF4 transcript levels enhances the responsiveness of genes regulated by N. The productivity increases seen were primarily due to the maintenance of photosystem II operating efficiency and carbon assimilation in plants when grown under limiting N conditions and not an overall increase in photosynthesis capacity. The increased biomass production and yield per plant in TaDWF4 OE lines could be linked to modified carbon partitioning and changes in expression pattern of the growth regulator Target Of Rapamycin, offering a route towards breeding for sustained yield and lower N inputs.

plant biology↗