Targeted modification of storage protein content resulting in improved amino acid composition of barley grain

Abstract

C-hordein in barley and ω-gliadins in wheat are members of the prolamins protein families. Prolamins are the major component of cereal storage proteins and composed of non-essential amino acids (AA) such as proline and glutamine therefore have low nutritional value. Using double stranded RNAi silencing technology directed towards C-hordein we obtained transgenic barley lines with up to 94.7% reduction in the levels of C-hordein protein relative to the parental line. The composition of the prolamin fraction of the barley parental line cv. Golden Promise was resolved using SDS-PAGE electrophoresis, the protein band were excised and the proteins identified by quadrupole-time-of-flight mass spectrometry. Subsequent SDS-PAGE separation and analysis of the prolamin fraction of the transgenic lines revealed a reduction in the amounts of C-hordeins and increases in the content of other hordein family members. Analysis of the AA composition of the transgenic lines showed that the level of essential amino acids increased with a concomitant reduction in proline and glutamine. Both the barley C-hordein and wheat ω-gliadin genes proved successful for RNAi-gene mediated suppression of barley C-hordein level. All transgenic lines that exhibited a reduction for C-hordein showed off-target effects: the lines exhibited increased level of B/γ-hordein while D-hordein level was reduced. Furthermore, the multicopy insertions correlated negatively with silencing.

Distribution of β-Glucan, Phenolic Acids, and Proteins as Functional Phytonutrients of Hull-Less Barley Grain

“…Edney et al [45] found that the concentrations of arginine, cysteine, isoleucine, lysine, methionine, and threonine were consistently higher in the hull-less barley samples than levels stated in the literature for barley. Sikdar et al [46], using double-stranded RNAi silencing technology directed towards C-hordeins, obtained transgenic barley lines with up to 94.7% reduction of C-hordeins as a storage protein with the lowest nutritional quality, what may be a promising approach for improving the nutritional value of barley.…”

High-throughput analysis of amino acids in plant materials by single quadrupole mass spectrometry

“…Here, not only information about the composition of essential amino acids is required but also that of non-essential amino acids as a proper balance is required to improve the utilization of the proteins in the diet [ 1 ]. The current societal focus on biorefining and bio-based economy has prompted a booming interest and spurred new research activities on how to exploit proteins extracted from green biomass [ 2 – 4 ]. However, these studies have often been performed without assessing the nutritional values of the obtained proteins by amino acid analysis [ 5 , 6 ].…”

Recent advances and perspectives in crop biofortification

“…Aminoacids, especially essential, such as lysine, methionine, threonine, and phenylalanine, have also been subjects of biofortification. Sikdar et al (2016) used RNAi technology to reduce a storage C-hordein protein by up to 97 %. Concomitantly, a drop in C-hordein content is balanced by an increase in B-hordein.…”

“…Attempts to biofortify crops have been conducted in order to meet demands of the mineral and vitamins for humans in populations under conditions of limited food sources. Prominent crops that have been subjected to biofortification efforts in recent years are: wheat (Singh et al 2017), barley (Sikdar et al 2016), maize (Zhu et al 2008, Du et al 2016, rice (Johnson et al 2011, Lee et al 2011, Ma et al 2015, sorghum (Che et al 2016), and banana (Paul et al 2017); all serving as staple food sources in large areas of the world. Microelements, vitamins, and fatty acids (FAs) constitute the main groups of substances that were subjected to biofortification (Table 1).…”