18 results about "Cell-free system" patented technology

The application provides an in-vitro cell-free protein synthesis system and kit containing exogenous magnesium ions and application thereof, including a D2P system (DNA-to-Protein system) and an mR2P system (mRNA-to-Protein system), and belongs to the technical field of protein synthesis. The in-vitro cell-free protein synthesis system containing exogenous magnesium ions adopts magnesium aspartate as a novel magnesium ion source, and especially in a eukaryotic cell-free system, compared with a traditional magnesium ion source, can significantly improve the protein synthesis efficiency and protein expression amount. A more efficient and higher-throughput in-vitro protein synthesis kit and a synthesis method of exogenous proteins are also provided, and the kit has the advantages of simplicity, convenience and low cost.

Compositions and methods disclosed herein relate to improved in vitro cell-free systems for various biological reactions. In one aspect, a composition of the present disclosure includes: a cell-free extract derived from an organism; a nucleic acid; and an organelle that is heterologous to the organism.

The disclosure provides biosynthetic platforms that generate olivetolic acid and its analogues at high titers from microbes, and in cell free systems.

This disclosure relates to single-molecule tracking methods, including high-throughput methods, for analyzing molecules in cell-free samples. The disclosure further provides a system for carrying out such methods.

Provided herein are membrane encapsulated cell-free systems comprising transcriptionally regulated biosensors for detecting analytes, and methods of their use. In embodiments, the biosensor is a riboswitch for detection of fluoride in a sample.

The application discloses a paper-based arsenic ion synthetic biosensor based on a cell-free system and application thereof, and comprises a cell-free arsenic ion sensing module and a semi-quantitative paper-based detection chip. The cell-free arsenic ion sensing module is composed of a synthetic arsenate reduction module, an arsenic ion specificity sensing module, a high-gain transcription amplification module and a visual reporting module connected in sequence. The synthetic arsenate reduction module converts pentavalent arsenic into trivalent arsenic; the arsenic ion specificity sensing module recognizes trivalent arsenic and removes transcription inhibition; the high-gain transcription amplification module amplifies the signal through ecf11 cascade; and the visual reporting module outputs a colorimetric or fluorescent signal. The application integrates an engineered promoter and an ecf11 cascade amplification system in a cell-free system, greatly improves sensitivity and dynamic range and keeps low leakage. The paper-based chip can present different pattern arrangements according to arsenic concentration and is easy to read. The application provides an environmentally friendly, portable and easy-to-read innovative method for monitoring arsenic pollution in drinking water.

The present disclosure relates to single molecule tracking methods, including high throughput methods, for analyzing molecules in cell-free samples. The present disclosure further provides systems for performing such methods.

The present invention relates to a cell lysate wherein the lysis buffer comprises magnesium acetate, the phosphorylation inhibitor K3L, the agonist of dephosphorylation GADD34 and a caspase inhibitor for use in, or as, a cell-free system and a method for preparing a cell-free protein synthesis system. Further provided is a method for expressing a protein in the cell-free protein synthesis system, a method for screening for a candidate compound for an effect on a target molecule and a use of the cell-free protein synthesis system. The present invention also provides a kit for the above-mentioned cell-free systems.

The invention relates to a method for preparing scDNA, which comprises the following steps: providing a circular nucleic acid template which is double-stranded and comprises a to-be-recombined ring-forming region and a to-be-enzyme-digested incision site arranged outside the region, and the to-be-recombined ring-forming region comprises an expression framework and a recombination site pair arranged on two wings of the expression framework; the present invention relates to a method for preparing a cyclic nucleic acid template, the cyclic nucleic acid template being amplified by RCA rolling circle amplification to form a concatemer, the concatemer comprising a tandem repeat unit of a cyclic nucleic acid template sequence, the concatemer being linearized by enzyme digestion in the incision site of the tandem repeat unit with a restriction enzyme simultaneously with or subsequent to the rolling circle amplification, the linearized concatemer comprising a cyclic nucleic acid template sequence, the cyclic nucleic acid template comprising a cyclic nucleic acid template sequence, and the cyclic nucleic acid template sequence comprising a cyclic nucleic acid template sequence. Free linearized nucleic acid is obtained; in a cell-free system, the linearized nucleic acid is recombined with a recombinase into scDNA comprising the expression framework. The method provided by the invention really realizes efficient preparation of scDNA.

The invention provides a cell-free toxin synthesis system and method. The system at least comprises: (1) a cell-free system; (2) an amplification system containing amplified plasmids of the cell-free system is added, the plasmids are plasmids containing coding toxin protein, the amplification system containing the amplified plasmids is obtained after the plasmids are added into the amplification system according to the preset concentration to be amplified, the preset concentration is 2-10 ng / mu L, and the preset concentration is not 2 ng / mu L. According to the method, plasmids with a certain concentration are added into a cell-free reaction system, so that the in-vitro expression efficiency of toxins is effectively improved, and particularly, the purification efficiency is more obviously improved.

The present disclosure relates to multi-enzyme pathways for the in-situ generation of the cofactor NADP from cost-effective reagents such as nicotinate, NMN (nicotinamide mononucleotide) and NAD (nicotinamide adenine dinucleotide) and the use of these pathways in NADP-dependent cell-free systems for the bio-based production of chemicals such as isobutanol.

The present invention relates to preparation of key drug intermediate, L-2-amino butyric acid (L-2-ABA) by a method of cell free system and biotransformation using genetically engineered strains from easily available economic substrates like citramalate or citraconate and enzymes like LeuCD, LeuB and ValDH or IlvE.

The present disclosure relates to single molecule tracking methods, including high-throughput methods, for analyzing molecules in cell-free samples. The present disclosure also provides systems for performing such methods.

The invention discloses a preparation method of tomato leaf vacuole-free cytoplasm. The method comprises the following steps: cutting tomato leaves, putting the cut tomato leaves into an enzymolysis buffer solution containing cellulase, macerozyme, mannitol, potassium chloride, MES, calcium chloride, beta-mercaptoethanol and fetal calf serum, and carrying out vacuumizing enzymolysis to obtain protoplast; removing vacuoles through density gradient centrifugation, and collecting tiny protoplasts; and performing staged homogenate cracking and centrifugal purification to obtain the tomato leaf vacuole-free cytoplasm. The vacuole-free cytoplasm prepared by the invention has dsRNA cleavage activity, and can be used for tomato virus in-vitro research and plant cell-free system research and development.

The invention belongs to the technical field of biomedicine, and particularly relates to a KV1.3 expression vector, a cell-free expression system, a recombination channel and application thereof. The recombinant expression vector is obtained by inserting a DNA (deoxyribonucleic acid) sequence as shown in SEQ ID NO.1 between an Nde I site and an Xho I site of a pIVEX-2. 4d vector, and the recombinant expression vector is named as pIVEX-K. The cell-free expression system is obtained by transferring the expression vector into a cell-free system. The recombination channel is obtained by recombining a cell-free expression system product and lipidosome. The recombinant channel can be applied to high-throughput screening of compounds influencing the potassium ion channel function, the influence of the compounds on the potassium channel function is directly displayed by monitoring the change of a fluorescence signal before and after a to-be-detected object is added, and the recombinant channel has the characteristics of simplicity in operation, short test period and high sensitivity.

The present technology relates to cell-free systems, methods, and kits for expressing proteins in vitro and evaluating the expressed proteins. In particular, the technology relates to cell-free systems, methods, and kits for expressing antibodies, antigen-binding fragment thereof, and antibody derivatives in vitro and evaluating the expressed antibodies, antigen-binding fragment thereof, and antibody derivatives.

Provided is a cell free system, a whole-cell biomanufacturing system, or methods which comprise a polypeptide that encodes an aldehyde dehydrogenase that directly reduces carboxylic acid containing compounds to aldehyde containing compounds without ATP or CoA-based activation. Also provided are additional polypeptides that can convert aldehyde containing compounds into additional functional groups and which can recycle nicotinamide-based coenzymes.