Abstract: An expression vector for expressing a target polypeptide in a prokaryotic cell is provided. The vector comprises a promoter operably linked to a polynucleotide encoding the target polypeptide operably linked to a eukaryotic secretion leader sequence, the eukaryotic secretion leader sequence encoding a signal peptide sequence selected from the group consisting of: a) MLKRSSWLATLGLLTVASVSTIVYA; b) MKKATFITCLLAVLLVSNPIWNA; c) MKVSAAALAVILIATALCAPASA; d) MKVSTAFLCLLLTVSAFSAQVLA; and e) MKCLLLALGLALACAAQA. Processes for expressing polypeptides and prokaryotic microorganisms comprising such vectors are also provided.

Abstract: An expression vector for expressing a target polypeptide in a prokaryotic cell is provided. The vector comprises a promoter operably linked to a polynucleotide encoding the target polypeptide operably linked to a eukaryotic secretion leader sequence, the eukaryotic secretion leader sequence encoding a signal peptide sequence selected from the group consisting of: a) MLKRSSWLATLGLLTVASVSTIVYA; b) MKKATFITCLLAVLLVSNPIWNA; c) MKVSAAALAVILIATALCAPASA; d) MKVSTAFLCLLLTVSAFSAQVLA; and e) MKCLLLALGLALACAAQA. Processes for expressing polypeptides and prokaryotic microorganisms comprising such vectors are also provided.

Abstract: A recombinant yeast of the genus Komagataella which has been transformed with a galactose promoter is provided. The yeast is preferably Komagataella phaffii, and the promoter is preferably a S. cerevisiae GAL1 or GAL10 promoter. Vectors and methods for producing recombinant polypeptides are also provided.

Abstract: A process for the production of a target recombinant polypeptide is provided. The process comprises expressing an expression cassette encoding the target polypeptide and co-expressing an expression cassette encoding a mitochondrial foldase. Preferred mitochondrial foldases include Mia 40 and Erv1, and homologs thereof.

Abstract: A process for the extraction of pDNA from cells is provided. In one aspect, the process comprises heating a liquid comprising the cells to an average temperature of from 95° C. to about 120° C. for a time of less than 10 seconds. In certain preferred aspects, the pDNA is extracted by the use of flow-through apparatus.

Abstract: A recombinant yeast of the genus Komagataella which has been transformed with a galactose promoter is provided. The yeast is preferably Komagataella phaffii, and the promoter is preferably a S. cerevisiae GAL1 or GAL10 promoter. Vectors and methods for producing recombinant polypeptides are also provided.

Abstract: A perfect palindrome operator sequence-based protein expression system is provided. The expression system comprises a promoter; and a perfect palindrome operator sequence, wherein the promoter is not T7. The expression system is preferably employed for the production of recombinant proteins by fermentation.

Abstract: A process for cell lysis is provided. The process comprises passing a mixture comprising a suspension of cells and a lysis reagent through a flow-through reactor, wherein the mixture passes through the reactor with pulsed or superimposed oscillating flow. The process is preferably employed for the preparation of biological products, such as pDNA or inclusion bodies.

Abstract: A process for the production of a target recombinant polypeptide is provided. The process comprises expressing a vector comprising a promoter operatively linked to an expression cassette for the target recombinant polypeptide; and a perfect palindrome operator sequence in a culture medium, wherein the culture medium is substantially free from antibiotic.

Abstract: A perfect palindrome operator sequence-based protein expression system is provided. The expression system comprises a promoter; and a perfect palindrome operator sequence, wherein the promoter is not T7. The expression system is preferably employed for the production of recombinant proteins by fermentation.

Abstract: A protein expression system is provided. The system comprises: a) a T7 RNA polymerase-dependent promoter operably linked to an expression cassette for a protein of interest; and b) an expression cassette for T7 RNA polymerase operably linked to a ?pL promoter and at least two perfect palindrome operator sequences.