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Summary Maize ARGOS8 is a negative regulator of ethylene responses. A previous study has shown that transgenic plants constitutively overexpressing ARGOS8 have reduced ethylene sensitivity and improved grain yield under drought stress conditions. To explore the targeted use of ARGOS8 native expression variation in drought‐tolerant breeding, a diverse set of over 400 maize inbreds was examined for ARGOS8 mRNA expression, but the expression levels in all lines were less than that created in the original ARGOS8 transgenic events. We then employed a CRISPR‐Cas‐enabled advanced breeding technology to generate novel variants of ARGOS8. The native maize GOS2 promoter, which confers a moderate level of constitutive expression, was inserted into the 5′‐untranslated region of the native ARGOS8 gene or was used to replace the native promoter of ARGOS8. Precise genomic DNA modification at the ARGOS8 locus was verified by PCR and sequencing. The ARGOS8 variants had elevated levels of ARGOS8 transcripts relative to the native allele and these transcripts were detectable in all the tissues tested, which was the expected results using the GOS2 promoter. A field study showed that compared to the WT, the ARGOS8 variants increased grain yield by five bushels per acre under flowering stress conditions and had no yield loss under well‐watered conditions. These results demonstrate the utility of the CRISPR‐Cas9 system in generating novel allelic variation for breeding drought‐tolerant crops.

Phytic acid in cereal grains and oilseeds is poorly digested by monogastric animals and negatively affects animal nutrition and the environment. However, breeding programs involving mutants with less phytic acid and more inorganic phosphate (P i ) have been frustrated by undesirable agronomic characteristics associated with the phytic acid-reducing mutations. We show that maize lpa1 mutants are defective in a multidrug resistance-associated protein (MRP) ATP-binding cassette (ABC) transporter that is expressed most highly in embryos, but also in immature endosperm, germinating seed and vegetative tissues. Silencing expression of this transporter in an embryo-specific manner produced low-phytic-acid, high-P i transgenic maize seeds that germinate normally and do not show any significant reduction in seed dry weight. This dominant transgenic approach obviates the need for incorporating recessive lpa1 mutations to create maize hybrids with reduced phytic acid. Suppressing the homologous soybean MRP gene also generated low-phytic-acid seed, suggesting that the strategy might be feasible for many crops.

Maize seeds are the major ingredient of commercial pig and poultry feed. Phosphorus in maize seeds exists predominately in the form of phytate. Phytate phosphorus is not available to monogastric animals and phosphate supplementation is required for optimal animal growth. Undigested phytate in animal manure is considered a major source of phosphorus pollution to the environment from agricultural production. Microbial phytase produced by fermentation as a feed additive is widely used to manage the nutritional and environmental problems caused by phytate, but the approach is associated with production costs for the enzyme and requirement of special cares in feed processing and diet formulation. An alternative approach would be to produce plant seeds that contain high phytase activities. We have over-expressed Aspergillus niger phyA2 gene in maize seeds using a construct driven by the maize embryo-specific globulin-1 promoter. Low-copy-number transgenic lines with simple integration patterns were identified. Western-blot analysis showed that the maize-expressed phytase protein was smaller than that expressed in yeast, apparently due to different glycosylation. Phytase activity in transgenic maize seeds reached approximately 2,200 units per kg seed, about a 50-fold increase compared to non-transgenic maize seeds. The phytase expression was stable across four generations. The transgenic seeds germinated normally. Our results show that the phytase expression lines can be used for development of new maize hybrids to improve phosphorus availability and reduce the impact of animal production on the environment.

REBCO (Rare-earth-based barium copper oxide) conductors are appropriate materials for high-field magnet applications. Vacuum impregnation using epoxy resin is a technique widely used for stable operation of superconducting coils. However, epoxy-impregnated REBCO coils often experience critical current degradation problems. Finding a suitable impregnating material for REBCO coils is important for their application in high-field superconducting magnets. A new toughness epoxy, IR-3, was developed recently. An in-depth understanding of IR-3 on the performance of REBCO coils is critically necessary for its application. Thus, this paper explores the effects of IR-3 impregnation on the performance of REBCO coils at 77 K and 4.2 K. The test results are compared to similar coils impregnated with CTD-101 K and MY750. Meanwhile, the radial stresses at 77 K in self-field and 4.2 K under 10 T were simulated. All epoxy impregnated REBCO coils showed no decay in critical current after thermal cycles at 77 K. When charged at 4.2 K in external fields of 5 T and 10 T, the IR-3 impregnated REBCO coils avoided performance degradation problems and had superior electrical stabilities. Combing the excellent performance at low temperatures, IR-3 is a promising candidate material for impregnating high-field REBCO coils.

Reduced phytic acid content in seeds is a desired goal for genetic improvement in several crops. Low-phytic acid mutants have been used in genetic breeding, but it is not known what genes are responsible for the low-phytic acid phenotype. Using a reverse genetics approach, we found that the maize (Zea mays) low-phytic acid lpa2 mutant is caused by mutation in an inositol phosphate kinase gene. The maize inositol phosphate kinase (ZmIpk) gene was identified through sequence comparison with human and Arabidopsis Ins(1,3,4)P3 5/6-kinase genes. The purified recombinant ZmIpk protein has kinase activity on several inositol polyphosphates, including Ins(1,3,4)P3, Ins(3,5,6)P3, Ins(3,4,5,6)P4, and Ins(1,2,5,6)P4. The ZmIpk mRNA is expressed in the embryo, the organ where phytic acid accumulates in maize seeds. The ZmIpk Mutator insertion mutants were identified from a Mutator F2 family. In the ZmIpk Mu insertion mutants, seed phytic acid content is reduced approximately 30%, and inorganic phosphate is increased about 3-fold. The mutants also accumulate myo-inositol and inositol phosphates as in the lpa2 mutant. Allelic tests showed that the ZmIpk Mu insertion mutants are allelic to the lpa2. Southern-blot analysis, cloning, and sequencing of the ZmIpk gene from lpa2 revealed that the lpa2-1 allele is caused by the genomic sequence rearrangement in the ZmIpk locus and the lpa2-2 allele has a nucleotide mutation that generated a stop codon in the N-terminal region of the ZmIpk open reading frame. These results provide evidence that ZmIpk is one of the kinases responsible for phytic acid biosynthesis in developing maize seeds.

Lack of sufficient water is a major limiting factor to crop production worldwide, and the development of drought-tolerant germplasm is needed to improve crop productivity. The phytohormone ethylene modulates plant growth and development as well as plant response to abiotic stress. Recent research has shown that modifying ethylene biosynthesis and signaling can enhance plant drought tolerance. Here, we report novel negative regulators of ethylene signal transduction in Arabidopsis (Arabidopsis thaliana) and maize (Zea mays). These regulators are encoded by the ARGOS gene family. In Arabidopsis, overexpression of maizeARGOS1(ZmARGOS1),ZmARGOS8, ArabidopsisARGOShomologORGAN SIZE RELATED1(AtOSR1), andAtOSR2reduced plant sensitivity to ethylene, leading to enhanced drought tolerance. RNA profiling and genetic analysis suggested that theZmARGOS1transgene acts between an ethylene receptor and CONSTITUTIVE TRIPLE RESPONSE1 in the ethylene signaling pathway, affecting ethylene perception or the early stages of ethylene signaling. Overexpressed ZmARGOS1 is localized to the endoplasmic reticulum and Golgi membrane, where the ethylene receptors and the ethylene signaling protein ETHYLENE-INSENSITIVE2 and REVERSION-TO-ETHYLENE SENSITIVITY1 reside. In transgenic maize plants, overexpression of ARGOS genes also reduces ethylene sensitivity. Moreover, field testing showed that UBIQUITIN1:ZmARGOS8maize events had a greater grain yield than nontransgenic controls under both drought stress and well-watered conditions.

The phytohormone ethylene regulates plant growth and development as well as plant response to environmental cues. ARGOS genes reduce plant sensitivity to ethylene when overexpressed in transgenic Arabidopsis (Arabidopsis thaliana) and maize (Zea mays). A previous genetic study suggested that the endoplasmic reticulum and Golgi-localized maize ARGOS1 targets the ethylene signal transduction components at or upstream of CONSTITUTIVE TRIPLE RESPONSE1, but the mechanism of ARGOS modulating ethylene signaling is unknown. Here, we demonstrate in Arabidopsis that ZmARGOS1, as well as the Arabidopsis ARGOS homolog ORGAN SIZE RELATED1, physically interacts with Arabidopsis REVERSION-TO-ETHYLENE SENSITIVITY1 (RTE1), an ethylene receptor interacting protein that regulates the activity of ETHYLENE RESPONSE1. The protein-protein interaction was also detected with the yeast split-ubiquitin two-hybrid system. Using the same yeast assay, we found that maize RTE1 homolog REVERSION-TO-ETHYLENE SENSITIVITY1 LIKE4 (ZmRTL4) and ZmRTL2 also interact with maize and Arabidopsis ARGOS proteins. Like AtRTE1 in Arabidopsis, ZmRTL4 and ZmRTL2 reduce ethylene responses when overexpressed in maize, indicating a similar mechanism for ARGOS regulating ethylene signaling in maize. A polypeptide fragment derived from ZmARGOS8, consisting of a Pro-rich motif flanked by two transmembrane helices that are conserved among members of the ARGOS family, can interact with AtRTE1 and maize RTL proteins in Arabidopsis. The conserved domain is necessary and sufficient to reduce ethylene sensitivity in Arabidopsis and maize. Overall, these results suggest a physical association between ARGOS and the ethylene receptor signaling complex via AtRTE1 and maize RTL proteins, supporting a role for ARGOS in regulating ethylene perception and the early steps of signal transduction in Arabidopsis and maize.

The operation status of the high voltage disconnector directly affects the safe and stable operation of the power system. This article uses the wireless frequency hopping communication technology of the communication module to achieve the temperature acquisition of the switch contacts and high voltage bus, to introduce the current value of the loop in ECS, and judge the operation status of the disconnector by considering the ambient temperature, calculating the temperature rise; And through the acquisition of the current of drive motor in the process of switch closing and opening, and fault diagnosis of the disconnector by analyzing the change rule of the drive motor current, the condition monitoring of the high voltage disconnector is realized.

High-throughput/low-cost/low-tech methods for phytic acid determination that are sufficiently accurate and reproducible would be of value in plant genetics, crop breeding and in the food and feed industries. Variants of two candidate methods, those described by Vaintraub and Lapteva (Anal Biochem 175:227–24, 1988 ; “VL” methods) and Huang and Lantzsch (J Sci Food Agric 34:1423–1426, 1983 ; “HL” methods), were evaluated. The primary concern with these methods is that, due to interference of matrix constituents including inorganic P, they can overestimate phytic acid and are ineffective at low levels of phytic acid. Twelve seed flours, representing lines of soybean, maize, barley and dry bean, containing a wide range of phytic acid levels, were analyzed by a minimum of eight cooperating laboratories using three variants of the VL method and two variants of the HL method. No method had consistently acceptable (˂2.0”) “Horwitz ratios”, a measure of reproducibility, although some treatments approached that. For example, one variant of the VL method when used to assay a soybean flour with a “standard” level of phytic acid had a Horwitz ratio of 2.15. Some variants of the VL method were adequate for analyses of cereal grains regardless of phytic acid level but none accurately measured phytic acid when at low levels in soybean flours. One variant of the HL method in which the 0.2 N HCl extraction media is modified to contain 10% Na 2 SO 4 , did accurately measure phytic acid levels in both cereal and legume flours regardless of endogenous phytic acid levels or matrix constituents.