A cofactor self-sufficient Escherichia coli and its construction method and application in the synthesis of L-glufosinate are provided. The present invention expresses a NADH kinase and key enzymes of the cofactor synthesis pathway in E. coli, and knocks out the genes of enzymes that catabolizes cofactor, and with the addition of co-metabolic intermediates during cell incubation, the intracellular NADP(H) concentration is increased by at least 50% and the catalytic activity of glutamate dehydrogenase by 2-fold, resulting in a significant increase in the spatiotemporal yield of the-glufosinate synthesis reaction.

The invention provides a genetically modified eukaryotic microorganism for anaerobic production of essential amino acids and optionally the co-production of one or more co-products. The microorganism is genetically modified to redirect carbon flow from PEP via oxaloacetate and asparatate semialdehyde, towards the synthesis of increased amounts of essential amino acids. The microorganism may be genetically modified to produce increased amounts of one or more co-product by enhancing carbon flow from PEP via pyruvate, acetyl CoA and malonyl CoA to produce alcohols and lipids, such as triglycerides, fatty esters, fatty alcohols, fatty aldehydes, fatty amides. The invention provides a method for anaerobic production of essential amino acids using the genetically modified eukaryotic microorganism and optionally co-production of said one or more co-products. The genetically modified eukaryotic microorganism may be used for the anaerobic production of essential amino acids and optionally the co-production of said one or more co-products.

A gene editing system of Candida viswanathii includes a Candida viswanathii, a first gene editing fragment and a second gene editing fragment. The first gene editing fragment successively includes a first homology arm and a screening gene. The second gene editing fragment is connected to a C-terminus of the first gene editing fragment and includes a second homology arm, a Cas9 expression cassette and a sgRNA cassette. The Cas9 expression cassette successively includes a Cas9 promoter, a Cas9 gene and three nuclear localization sequences. The sgRNA cassette successively includes a sgRNA promoter, a first ribozyme, a targeting sequence, a scaffold and a second ribozyme. The first gene editing fragment and the second gene editing fragment are constructed as a linear fragment for gene editing of a chromosome of the Candida viswanathii.

A cofactor self-sufficient Escherichia coli and its construction method and application in the synthesis of L-glufosinate are provided. The present invention expresses a NADH kinase and key enzymes of the cofactor synthesis pathway in E. coli, and knocks out the genes of enzymes that catabolizes cofactor, and with the addition of co-metabolic intermediates during cell incubation, the intracellular NADP(H) concentration is increased by at least 50% and the catalytic activity of glufosinate dehydrogenase by 2-fold, resulting in a significant increase in the spatiotemporal yield of the glufosinate synthesis reaction.

Provided is a genetically engineered yeast cell having increased NADPH production, a method of increasing a NADPH level in a yeast cell, a method of preparing the genetically engineered yeast cell, and a method of producing lactate using the genetically engineered yeast cell.

A gene editing system of Candida viswanathii includes a Candida viswanathii, a first gene editing fragment and a second gene editing fragment. The first gene editing fragment successively includes a first homology arm and a screening gene. The second gene editing fragment is connected to a C-terminus of the first gene editing fragment and includes a second homology arm, a Cas9 expression cassette and a sgRNA cassette. The Cas9 expression cassette successively includes a Cas9 promoter, a Cas9 gene and three nuclear localization sequences. The sgRNA cassette successively includes a sgRNA promoter, a first ribozyme, a targeting sequence, a scaffold and a second ribozyme. The first gene editing fragment and the second gene editing fragment are constructed as a linear fragment for gene editing of a chromosome of the Candida viswanathii.

The invention provides a recombinant Yarrowia lipolytica. The recombinant Yarrowia lipolytica comprises a heterologous polynucleotide encoding a fusion protein. The fusion protein comprises a first amino acid sequence and a second amino acid sequence. The first amino acid may be homologous to the amino acid sequence of 3-ketoacyl CoA thiolase (3KAT), The second amino acid sequence may be homologous to the amino acid sequence of a fatty acyl-CoA reductase (FAR). Also provided is a method for producing one or more fatty alcohols by the recombinant Yarrowia lipolytica and a method for preparing the recombinant Yarrowia lipolytica.