139 results about "Cell-free protein synthesis" patented technology

Various nucleic acid-based matrixes are provided, comprising nucleic acid monomers as building blocks, as well as nucleic acids encoding proteins, so as to produce novel biomaterials. Methods of utilizing such biomaterials include delivery of biologically active agents, cell and tissue culture, and cell-free protein synthesis.

The invention provides a theoretical design and technical design of cell-free protein synthesis for DNA replication, transcription and translation coupling, a preparation, a kit and a preparation method. Specifically, with the application of the in-vitro cell-free synthesis system provided by the invention, complex protein can be synthesized, and moreover, DNA and mRNA can be synthesized; and effective, high-throughput and quite convenient protein synthesis can be completed with the use of a DNA template by a minute quantity (nanogram-microgram).

It is intended to provide a process for producing a plant germ extract wherein the step of breaking a plant germ into fine pieces is carried out by impacting or cutting and/or in the presence of an extraction solvent, and a germ extract obtained by this process which is contaminated with little impurities unnecessary or exerting undesirable effects in cell-free protein synthesis. Thus, cell-free protein synthesis can be performed at a high stability and a high efficiency.

The present invention provides a preparation method of a yeast extract solution for cell-free protein synthesis, which solution is easy to prepare and is capable of synthesizing a higher amount of a protein than by conventional yeast extract solutions, the yeast extract solution, a cell-free synthesis method of protein, which uses the yeast extract solution, and a kit for cell-free protein synthesis containing the yeast extract solution. The method of the present invention includes rupturing a yeast cell in a frozen state, and obtaining an extract thereof.

The present invention provides a method for monitoring of profile changes of components in a dynamic system such as a cell-free in vitro protein synthesis system by using liquid chromatography (LC) combined with mass spectrometry (MS). In an additional aspect, this invention provides a method for enhancing the yield and/or reproducibility in a cell-free protein synthesis system by modulating the level and/or activity of a protein component that has regulatory effects on the system.

Compositions and methods are provided for the in vitro synthesis of biological molecules in reaction mixtures comprising anti-foam agents. The reaction mix comprising antifoam agent may be a scaled up reaction, e.g. in reaction volume greater than at least about 15 ul. Reactions may be performed in various reactors, as known in the art, which include stirred reactors, bubble-column reactors; and the like.

The present invention is intended to provide a safe and quick means for screening a drug to a bioactive protein, in particular, an inhibitor, using a cell-free protein synthesis system with the use of a wheat embryo extract solution. The present inventors have strenuously studied to solve the matters above and finally completed the present invention by, in a system with the use of a wheat embryo, among cell-free protein synthesis means, constructing a synthesis system of a bioactive protein while sustaining its activities, and constructing a system for screening an inhibitor candidate to SARS 3CL^pro, as an example using the synthesis system.

The invention discloses a method for expression of fatty acid desaturase by an acellular protein synthesis system, and belongs to the enzyme engineering field. The method uses the wheat germ acellular protein synthesis system for cloning and expression of omega 3 desaturase gene (FADS15) from mortierella alpine ATCC32222 (American type culture collection32222) without the need for preparation of mRNA, the expression amount is high, subsequent purification steps are simple, and the method lays the foundation for the next step of research of membrane protein crystal structures and functions.

The present invention provides a novel extract solution for cell-free protein synthesis, which contains at least an extract derived from a Bombyx mori L. tissue and an exogenous mRNA, which is easy to prepare and which is capable of realizing cell-free protein synthesis that can synthesize glycoprotein, a cell-free protein synthesis method using it, and a method for preparing an extract solution for cell-free protein synthesis, which includes adding an exogenous mRNA to an extract derived from a Bombyx mori L. tissue.

Prepared is an extract composition having an improved protein synthetic activity in a cell-free protein synthesis system using a mammalian cultured cell extract. An eukaryotic translation initiation factor and/or translational regulator are added to a cell-free protein synthesis system comprising an extract prepared from cultured mammalian cells and a template mRNA. These factors are one or more selected from the group consisting of eukaryotic translation initiation factors 4E (eIF4E), 2 (eIF2) and 2B (eIF2B), and eukaryotic translational regulator p97.

The present invention provides a wheat germ cell-free protein synthesis system for high level expression of recombination snake venom kininogenase. According to the system, snake venom kininogenase gene of agkistrodon halys pallas is cloned through reverse transcription, wheat germ cell-free expression plasmid pCS<2+>/VK is constructed, a wheat germ extract is prepared, an in vitro transcription and translation system is established, the snake venom kininogenase is expressed and the system is optimized. The N terminal of the exogenous protein expressed by the expression system has the His protein tag, and a Ni column elution method is adopted to carry out purification, such that the subsequent purification of the recombinant protein is substantially simplified, the purification efficiency is high, and the cost is low. The exogenous protein further has biological activity, and provides the foundation for the next clinical study.

Provided is a process for producing a protein with post-translational modification in a cell-free protein synthesis system having a higher protein synthetic activity and glycosylation capability. The process of the present invention comprises preparing a cell extract from cultured cells of an immortalized mammalian cell line that has enhanced protein secreting activity, and adding an mRNA encoding a glycoprotein to the cell extract.

This invention relates to methods of producing a rare amino acid- or non-natural amino acid-containing protein in a cell free protein synthesis system and kits for use in and for accomplishing same. Specifically, the methods comprise the steps of expressing at least one orthogonal suppressor tRNA (o-tRNA)/aminoacyl-tRNA synthetase (aaRS) pair or derivatives thereof specific for incorporation of a rare amino acid- or non-natural amino acid in an E. coli organism; preparing a lysate of said E. coli organism expressing said orthogonal suppressor tRNA (o-tRNA)/aminoacyl-tRNA synthetase (aaRS) pair; and contacting said lysate with a template DNA containing a mutant gene in which at least one amino acid codon at a given site of the protein-encoding gene has been mutated into an amber or ochre mutation and further providing a cognate rare amino acid or non-natural amino acid and other factors necessary for protein synthesis; wherein protein synthesis occurs following said contact to produce a protein containing said at least one rare amino acid or said non-natural amino acid. Kits for use are described, as well.

Methods and kits are provided for calibrating a cell-free protein synthesis reaction for optimal activity. The method includes the steps of providing an extract competent for cell-free protein synthesis (CFPS); performing cell-free protein synthesis with the extract; measuring a first reaction end-point where in vitro protein synthesis plateaus; measuring a second reaction end-point where Energy Charge of the extract declines to a level in a range from about 0.40 to about 0.80 of Energy Charge of a control extract; and adjusting Energy Charge of the extract to a level in a range from about 0.80 to about 1.0 of the control extract.

According to the present invention, a composition possessing cell-free protein synthesis activity with reduced contaminating lipopolysaccharide, and a method for producing a protein using the same are provided. When ribosome display is performed using the composition and method for protein production of the present invention, the background that is caused by non-specific binding is reduced, so that a nucleic acid that encodes the desired polypeptide can be selected with high accuracy and high efficiency.