SummaryFunctional stay‐green is a valuable trait that extends the photosynthetic period, increases source capacity and biomass and ultimately translates to higher grain yield. Selection for higher yields has increased stay‐green in modern maize hybrids. Here, we report a novel QTL controlling functional stay‐green that was discovered in a mapping population derived from the Illinois High Protein 1 (IHP1) and Illinois Low Protein 1 (ILP1) lines, which show very different rates of leaf senescence. This QTL was mapped to a single gene containing a NAC‐domain transcription factor that we named nac7. Transgenic maize lines where nac7 was down‐regulated by RNAi showed delayed senescence and increased both biomass and nitrogen accumulation in vegetative tissues, demonstrating NAC7 functions as a negative regulator of the stay‐green trait. More importantly, crosses between nac7 RNAi parents and two different elite inbred testers produced hybrids with prolonged stay‐green and increased grain yield by an average 0.29 megagram/hectare (4.6 bushel/acre), in 2 years of multi‐environment field trials. Subsequent RNAseq experiments, one employing nac7 RNAi leaves and the other using leaf protoplasts overexpressing Nac7, revealed an important role for NAC7 in regulating genes in photosynthesis, chlorophyll degradation and protein turnover pathways that each contribute to the functional stay‐green phenotype. We further determined the putative target of NAC7 and provided a logical extension for the role of NAC7 in regulating resource allocation from vegetative source to reproductive sink tissues. Collectively, our findings make a compelling case for NAC7 as a target for improving functional stay‐green and yields in maize and other crops.

Most of the maize kernel oil is located in the embryo while the majority of starch is located in the endosperm. Maize kernel composition and value are affected significantly by the ratio of the embryo size to the endosperm size; however, the genetic regulation of embryo to endosperm ratio (EER) in maize is unknown. Here we identified ZmGE2 gene, which encodes a cytochrome p450 protein, as a gene associated with EER variation in maize. We first expressed rice Giant Embryo (GE) gene driven by oleosin promoter in maize and detected a 23.2 % reduction in EER in transgenic seeds, demonstrating the existence of evolutionarily conserved mechanisms for EER determination in rice and maize. We next identified maize GE2, a homolog of rice GE sharing 70 % identity in amino sequence, as a candidate based on the similar expression pattern and co-localization with a previously detected QTL for EER. Followed by linkage and association mapping, a 247-bp transposable element (TE) insertion in 3'-untranslated region of ZmGE2 gene was identified to be associated with increase in EER and kernel oil content. Expression level of the favorable ZmGE2 allele containing the 247-bp TE insertion was strongly reduced. In addition, the 247-bp TE insertion site was a selection target during the artificial long-term selection for the high EER trait in a high oil population. This is the first report that demonstrates an association of ZmGE2 with EER variation in maize and identifies ZmGE2 gene as a promising target for manipulation of EER and grain composition by either transgenic approach or molecular breeding in maize.

The gene encoding acyl-CoA:diacylglycerol acyltransferase (DGAT1-2) is a key quantitative trait locus that controls oil content and oleic acid composition in maize kernels. Here we re-sequenced the DGAT1-2 region responsible for oil variation in a maize landrace set and in 155 inbred lines (35 high-oil and 120 normal lines). The high-oil DGAT1-2 allele was present in most Northern Flint and Southern Dent populations but was absent in five of eight Corn Belt Dent open-pollinated populations and in most of the earlier inbred lines. Loss of the high-oil DGAT1-2 allele possibly resulted from genetic drift in the early twentieth century when a few Corn Belt Dent populations were selected for the development of high-grain-yield inbred lines. Association analysis detected significant effects of two PCR-based functional markers (HO06 and DGAT04; developed based on DGAT1-2 polymorphisms) on kernel oil content and oleic acid composition using the 155 inbred lines. Zheng58 and Chang7-2, the parent inbred lines of elite hybrid Zhengdan958, were used to transfer the favorable allele from the high-oil line By804 using marker-assisted backcrossing with the two functional markers. In BC5F2:3 populations, oil content of the three genotypes (−/−, +/−, and +/+) was, respectively, 3.37, 4.20, and 4.61% (Zheng58 recipient line) and 4.14, 4.67, and 5.25% (Chang7-2 recipient line). Oil content of homozygous kernels containing the high-oil DGAT1-2 allele increased by 27–37% compared with recurrent parents. Hence, these functional markers can be used to re-introduce the high-oil DGAT1-2 allele into modern inbred lines for increased oil content through marker-assisted backcrossing.

Increasing seed oil production is a major goal for global agriculture to meet the strong demand for oil consumption by humans and for biodiesel production. Previous studies to increase oil synthesis in plants have focused mainly on manipulation of oil pathway genes. As an alternative to single-enzyme approaches, transcription factors provide an attractive solution for altering complex traits, with the caveat that transcription factors may face the challenge of undesirable pleiotropic effects. Here, we report that overexpression of maize (Zea mays) LEAFY COTYLEDON1 (ZmLEC1) increases seed oil by as much as 48% but reduces seed germination and leaf growth in maize. To uncouple oil increase from the undesirable agronomic traits, we identified a LEC1 downstream transcription factor, maize WRINKLED1 (ZmWRI1). Overexpression of ZmWRI1 results in an oil increase similar to overexpression of ZmLEC1 without affecting germination, seedling growth, or grain yield. These results emphasize the importance of field testing for developing a commercial high-oil product and highlight ZmWRI1 as a promising target for increasing oil production in crops.

ABSTRACTNear‐infrared reflectance (NIR) spectroscopy can be used for fast and reliable prediction of organic compounds in complex biological samples. We used a recently developed NIR spectroscopy instrument to predict starch, protein, oil, and weight of individual maize (Zea mays) seeds. The starch, protein, and oil calibrations have reliability equal or better to bulk grain NIR analyzers. We also show that the instrument can differentiate quantitative and qualitative seed composition mutants from normal siblings without a specific calibration for the constituent affected. The analyzer does not require a specific kernel orientation to predict composition or to differentiate mutants. The instrument collects a seed weight and a spectrum in 4–6 sec and can collect NIR data alone at a 20‐fold faster rate. The spectra are acquired while the kernel falls through a glass tube illuminated with broad spectrum light. These results show significant improvements over prior single‐kernel NIR systems, making this instrument a practical tool to collect quantitative seed phenotypes at high throughput. This technology has multiple applications for studying the genetic and physiological influences on seed traits.

Plant oil is an important renewable resource for biodiesel production and for dietary consumption by humans and livestock. Through genetic mapping of the oil trait in plants, studies have reported multiple quantitative trait loci (QTLs) with small effects, but the molecular basis of oil QTLs remains largely unknown 1 , 2 , 3 , 4 , 5 . Here we show that a high-oil QTL ( qHO6 ) affecting maize seed oil and oleic-acid contents encodes an acyl-CoA:diacylglycerol acyltransferase (DGAT1-2), which catalyzes the final step of oil synthesis. We further show that a phenylalanine insertion in DGAT1-2 at position 469 (F469) is responsible for the increased oil and oleic-acid contents. The DGAT1-2 allele with F469 is ancestral, whereas the allele without F469 is a more recent mutant selected by domestication or breeding. Ectopic expression of the high-oil DGAT1-2 allele increases oil and oleic-acid contents by up to 41% and 107%, respectively. This work provides insights into the molecular basis of natural variation of oil and oleic-acid contents in plants and highlights DGAT as a promising target for increasing oil and oleic-acid contents in other crops.