Tarlyn, N. | bioRxiv Science

bioRxiv2026

Bioengineering of Pea (Pisum sativum) for the Expression of Myoglobin, a Heme-containing Animal Protein

Myoglobin, an oxygen-binding animal protein, was engineered in Pisum sativum (pea) to explore its potential as a food ingredient and balance the amino acid profile. In this study, minimal expression cassettes and binary vectors were used to express bovine myoglobin using particle gun and Agrobacterium-mediated transformation, respectively. Successful integration and expression of the myoglobin gene was achieved in P. sativum, with both methods yielding similar transformation efficiencies (~1%). Expression analysis of T2 seeds revealed that Agrobacterium-mediated transformation-derived transgenic lines that expressed myoglobin under the regulation of a Soybean 7S seed-specific promoter and Tobacco Etch Virus (TEV) translation enhancer and a chimeric Rb7MAR Terminator (Ps-BpRG13 events) consistently yielded the highest level of expression (0.32-1.57% of TSP), while transgenic lines with myoglobin expression under the regulation of a Soybean Phaseolin promoter and Rb7MAR Terminator (Ps-BpRG14 events) resulted in moderate levels of heterologous protein expression (0.13-0.83% TSP). Transgenic events with constitutive 2xCaMV35S promoter, TEV translation enhancer and Rb7MAR terminator (Ps-BpRG15 events) exhibited the lowest level of myoglobin expression (0.09-0.14% TSP). Co-bombardment of two minimal expression cassettes - one with myoglobin under the regulation of the Phaseolin promoter and Rb7MAR Terminator and the other with the nptII selectable marker under the regulation of a 2X constitutive CaMV35S promoter, TEV translational enhancer and TNOS Terminator, yielded lines that exhibited variable expression (0.03-0.77% TSP), with some events comparable in expression to Agrobacterium-derived Ps-pRG14 events. To the best of our knowledge, this is the first report of producing a heme-containing animal protein, myoglobin, in peas, with potential implications for sustainable production of food ingredients and nutritionally fortified and value-added plant products using molecular farming.

plant biology↗

bioRxiv2026

Extensive Novel Genomic Variations in Mutant European Pear Individuals Revealed by Mapping to a Pangenome Reference

European pear (Pyrus communis) is the most widely cultivated Pyrus species outside of Asia; however, production has been declining in Europe, Oceania, and North America. Most European pear cultivars are over 100 years old and face pressure from disease, a changing climate, and challenging postharvest characteristics, while demand for increased production efficiency rises. To generate germplasm with desirable characteristics to address these concerns, a mutation breeding approach was chosen, with crosses made between four economically significant cultivars ( Bartlett, dAnjou, Abbe Fetel, and Comice) using gamma-irradiated pollen. This resulted in 49 viable offspring, of which 37 have survived at least 10 years. Nanopore whole-genome sequencing was used to test the success of this approach and screen for variants of interest. Sequence reads were mapped to both a lightweight, purpose-built pangenome derived from assemblies of parental haplotypes and a linear reference genome, enabling the high-quality discovery of variants of all sizes, ranging from single-base substitutions to megabase-scale deletions, with the overwhelming majority being small variants. The overall rate of mutation was 153 novel small variants and 0.228 novel structural variants per Gray of absorbed gamma radiation. Alternate ploidy levels were detected in four lines, which included three triploids and one tetraploid. While the resulting individuals appear incapable of floral development, they may be of utility as rootstock cultivars and a valuable genetic resource for understanding the underlying basis of structural traits.

genomics↗