Disclosed herein are methods of increasing nitrogen fixation in a non-leguminous plant. The methods can comprise exposing the plant to a plurality of bacteria. Each member of the plurality comprises one or more genetic variations introduced into one or more genes or non-coding polynucleotides of the bacteria's nitrogen fixation or assimilation genetic regulatory network, such that the bacteria are capable of fixing atmospheric nitrogen in the presence of exogenous nitrogen. The bacteria are not intergeneric microorganisms. Additionally, the bacteria, in planta, produce 1% or more of the fixed nitrogen in the plant.

An artificial non-coding RNA (ncRNA) module constructed by a synthetic biology technique and the use of the artificial ncRNA module in the construction of an artificial nitrogen fixation system are disclosed. The RNA module can enhance the post-transcriptional stability of nifHDK mRNA by interacting with a nitrogenase coding gene nifHDK mRNA, thereby improving the nitrogen fixation ability of a chassis microorganism. A fusion expression vector carrying the artificial RNA module is constructed and transformed into different chassis nitrogen-fixing microorganisms. It is confirmed through experiments that, under nitrogen fixation conditions, the artificial RNA module of the present disclosure can significantly improve the nitrogenase activity of a recombinant engineering bacterial strain.

Autotrophic nitrogen fixation genes and an autotrophic nitrogenase expression cassette for plants and uses thereof are disclosed. Nitrogenase related genes of nitrogen fixation gene cluster from Paenibacillus sp. are specifically optimized by plant expression patterns through synthetic biology, to obtain autotrophic nitrogen fixation genes, which are then constructed into plant constitutive expression units. The expression units are assembled in a plant expression vector by isocaudarner cloning method and transformed into plants, thereby realizing autotrophic nitrogen fixation of the plants.

Present invention discloses plants and plant cells comprising Streptomyces thermoautotrophicus nitrogenase and capable able of nitrogen fixation. Methods to generate said plants and plant cells are disclosed. This invention is instrumental for producing plants, including agriculturally important crops, with reduced or abolished requirements for nitrogen fertilizer.

The invention relates to a nano-biohybrid organism (or nanorg) comprising one of at least seven different core-shell quantum dots (QDs) or gold nanoparticle clusters, with excitations ranging from ultraviolet to near-infrared energies, couple with targeted enzyme sites in bacteria. When illuminated by light, these QDs drive the renewable production of biofuel molecules and chemicals using carbon-dioxide (CO.sub.2), water, and nitrogen (from air) as substrates. Nanorgs catalyze light-induced air-water-CO.sub.2 reduction with a high turnover number (TON) of approximately 10.sup.6-10.sup.8 (mols of product per mol of cells) to biofuels such as isopropanol (IPA), butane diol, gasoline additives, gasoline substitutes, 2,3-butanediol (BDO), C11-C15 methyl ketones (MKs), and hydrogen (H2); Sand chemicals such as formic acid (FA), ammonia (NH.sub.3), ethylene (C.sub.2H.sub.4), and degradable bioplastics, e.g. polyhydroxybutyrate (PHB). These nanorg cells function as nano-microbial factories powered by light.

The present invention relates to a method for promoting acetylglucosamine synthesis of Bacillus subtilis, which belongs to the field of genetic engineering. The present invention adopts the recombinant Bacillus subtilis BSGNKAP2 as a starting strain, exogenously introducing pyruvate carboxylase BalpycA derived from Bacillus cereus, eliminating the central carbon metabolism overflow of the Bacillus subtilis and avoiding the synthesis of the by-product acetoin; further, five exogenous reducing force metabolic reactions are introduced to replace the reaction of generating NADH in glycolysis pathway and tricarboxylic acid cycle to reconstruct intracellular reducing force metabolism, which specifically comprise glyceraldehyde-3-phosphate ferredoxin dehydrogenase, isocitrate NAD.sup.+ dehydrogenase, a malate quinone dehydrogenase, a ketoacid ferredoxin oxidoreductase and a nitrogenase ferritin. In a shake-flask fermentation process using a complex medium, acetylglucosamine yield of the recombinant strain BSGNKAP8 is 24.50 g/L, acetylglucosamine/glucose yield is 0.469 g/g, respectively 1.97 times and 2.13 times of those of the starting strain BSGNKAP2.

The present invention relates to methods and means for producing nitrogenase polypeptides in the mitochondria of plant cells.

Disclosed herein are methods of increasing nitrogen fixation in a non-leguminous plant. The methods can comprise exposing the plant to a plurality of bacteria. Each member of the plurality comprises one or more genetic variations introduced into one or more genes or non-coding polynucleotides of the bacteria's nitrogen fixation or assimilation genetic regulatory network, such that the bacteria are capable of fixing atmospheric nitrogen in the presence of exogenous nitrogen. The bacteria are not intergeneric microorganisms. Additionally, the bacteria, in planta, produce 1% or more of the fixed nitrogen in the plant.

The present disclosure provides engineered gram-positive microbes that are able to fix atmospheric nitrogen and deliver such to plants in a targeted, efficient, and environmentally sustainable manner. The utilization of the taught microbial products will enable farmers to realize more productive and predictable crop yields without the nutrient degradation, leaching, or toxic runoff associated with traditional synthetically derived nitrogen fertilizer.

Disclosed herein are biohybrid protein complexes capable of using light energy to photocatalyze the reduction of N.sub.2 into NH.sub.3. Also provided are methods of using biohybrid protein complexes to enzymatically reduce N.sub.2 to NH.sub.3 using light rather than chemical energy as the driving force. These methods may also include the production and isolation of ammonia, hydrogen or both.