53 results about "Translational efficiency" patented technology

The present invention relates to a pharmaceutical composition comprising a modified mRNA that is stabilised by sequence modifications and optimised for translation. The pharmaceutical composition according to the invention is particularly well suited for use as an inoculating agent, as well as a therapeutic agent for tissue regeneration. In addition, a process is described for determining sequence modifications that promote stabilisation and translational efficiency of modified mRNA of the invention.

It was the object of the present invention to provide RNA with increased stability and translation efficiency and means for obtaining such RNA. It should be possible to obtain increased grades of expression by using said RNA in gene therapy approaches.

Described herein are rules to modify natural mRNAs or to engineer synthetic mRNAs to increase their translation efficiencies. These rules describe modifications to mRNA coding and 3′ UTR sequences intended to enhance protein synthesis by: 1) decreasing ribosomal diversion via AUG or non-canonical initiation codons in coding sequences, and/or 2) by evading miRNA-mediated down-regulation by eliminating one or more miRNA binding sites in coding sequences.

The invention provides a multiple His sequence tag and application of the multiple His sequence tag to protein expression and purification. Specifically, a nucleic acid builder is composed of coding sequences of nHis, catenation sequences (including the codon optimized catenation sequence and the un-optimized catenation sequence) and coding sequences of foreign proteins. By applying the nucleic acid builder to a protein synthesis system, the translation efficiency of target proteins can be improved, and the foreign proteins can be expressed and purified.

The present invention discloses a method for modulating the production of a protein from a polynucleotide in a CHO cell by replacing at least one codon of the polynucleotide with a synonymous codon that has a higher or lower translation efficiency in the CHO cell than the codon it replaces, or by introducing into the CHO cell a polynucleotide that codes for an iso-tRNA which limits the rate of production of the polypeptide and which corresponds to a codon of the first polynucleotide. The present invention also discloses the use of a protein-encoding polynucleotide whose codon composition has been modified for enhanced production of the protein in CHO cells.

The present invention provides a cyclic RNA preferable for carrying out rotary protein translation in which translation domains other than that of the target protein are sufficiently short and translation efficiency is high, and a method for producing protein that uses this cyclic RNA as template. More specifically, the present invention provides a cyclic RNA that encodes a protein, has a full-length number of bases that is equal to or greater than 102 and is a multiple of 3, has at least one start codon, does not have a stop codon in the same reading frame as the start codon, and does not contain an internal ribosome entry site (IRES). In addition, the present invention provides a method for producing protein in a eukaryotic cell expression system that consists of using the aforementioned cyclic RNA as template and expressing a protein encoded by that cyclic RNA.

The present invention relates to a DNA fragment for improving translation efficiency, and a recombinant vector containing the same, and more specifically, to a DNA fragment which comprises any one nucleotide sequence selected from the group consisting of SEQ ID NOs: 1-6, SEQ ID NOs: 8-10, SEQ ID NOs: 13 and 14 and SEQ ID NO: 16, and improves the translation efficiency of a heterologous protein placed in the downstream, and a recombinant vector containing the DNA fragment. The DNA fragment for improving translation efficiency according to the present invention and a recombinant vector containing the same can improve the translation of a heterologous protein in a transgenic plant. In addition, if a leader polynucleotide inducing the targeting to a particular cellular organelle of a plant is further linked to the recombinant vector in an operable manner, the heterologous protein is targeted to the specific cellular organelle and can be stably accumulated, thereby enabling the mass production of the heterologous protein from the plant.

An objective of the present invention is to provide methods of synthesizing peptides containing structurally diverse amino acids using cell-free translation systems, which can accomplish excellent translational efficiency as compared to conventional techniques (the conventional techniques being methods which involve preparing aminoacyl-tRNAs which do not have protecting groups outside the translation systems without using ARS, and then adding the prepared aminoacyl-tRNAs into translation systems). In the present invention, it was found that amino acid-containing peptides can be synthesized efficiently by protecting an amino acid linked to tRNA with an appropriate protecting group, and then performing the step of deprotecting the protecting group of the amino acid linked to tRNA and the step of peptide translation from a template nucleic acid in a cell-free translation system in parallel.

When the initiation suppression method was used for translation of a peptide having at its N terminus an amino acid residue carrying a thiol group near its amino group with specific protecting groups being introduced to the thiol group and the amino group, it was found that not only the probability of initiation of amino acid translation reaction was improved, but also production of cleaved peptides was suppressed and translation efficiency and purity were improved. Furthermore, it was found that it is possible to efficiently promote the cyclization reaction of the peptide through amide bond formation. Based on these findings, the inventors discovered novel methods for preparing complexes between nucleic acids and peptides containing various unnatural amino acids and having an amide bond-mediated cyclized portion.

The invention discloses establishment of a self-activated Gal4/UAS system expression cassette capable of improving gene expression. The expression cassette with good improvement effect is obtained byoptimizing the number of UAS and the types of GAL 4 activation/binding domains; the expression cassette is started by the strong promoter CMV of the expression cassette and improves transcription andtranslation efficiency in virtue of a Gal4/UAS system, so the effects of automatic activation and cascade amplification are obtained; high improvement effect is maintained in 5 to 10 days after transfection, and highest expression activity is obtained in the eighth day, so it is proved that the expression cassette can improve the stability of gene expression; and improvement effect is about 9 times better in CHO cells specially producing recombinant proteins. Compared with conventional gene expression cassettes, the expression cassette provided by the invention is simple in structure and substantial in improvement effect, and provides an approach and novel method for non-virus transfection approaches of gene therapy and enhancement of the expression of target genes in production of recombinant proteins.

In order to solve the problems that code cross-platform translation efficiency is low, requirements for translators are high, and translation cost is increased, the invention provides a code translation method which comprises the steps that a first code is obtained, the first code is written based on a platform-dependent language of a source platform, then at least one translation rule matched with the first code is determined, and the first code is translated according to the at least one translation rule. The translation rule is used for directly translating a code of one platform into a code of another platform, and then translating the first code by using the translation rule to obtain a second code suitable for the target platform. According to the method, automatic cross-platform translation of the codes is realized through the translation rule, and the translation efficiency and the translation precision are improved.

The invention belongs to the technical field of molecular biology and particularly relates to a method for efficiently expressing a recombinant human fibroblast growth factor 21 (rhFGF21) gene by using bacillus subtilis. Firstly, the method adjusts the transcription and translation efficiency of a target protein gene by adding different types of cistron sequences to the 5' end of the target protein gene so as to optimize the soluble expression level of the protein gene, and the method can be applied to the optimal expression of different types of heterologous protein; secondly, the method further optimizes disulfide bond related genes to construct a vector to promote the disulfide bond folding efficiency of rhFGF21, overexpresses a molecular chaperone system to construct a vector suitablefor the folding efficiency increasing of rhFGF21 protein, and knocks out various extracellular protease genes to construct a bacillus subtilis gene engineering strain for improving the extracellular stability of rhFGF21; and finally, the method provides a bacillus subtilis gene engineering strain with good application prospect and capable of achieving efficient secretory expression of the rhFGF21protein.

The invention discloses a method for improving the stability of in-vitro synthetic mRNA. After PCR tailing, cap structure analogue modification and base analogue modification are carried out respectively with a luciferase mRNA synthesis template plasmid as a template, the mRNA is synthesized by using an MEGAscript T7 kit according to an instruction book under the action of T7 RNA polymerase. Through researches about the length of polyA tails and the translation efficiency and stability of cap structure analogues and base analogues on the in-vitro synthetic mRNA, it is found that through a series of modification, the translation efficiency of the in-vitro synthetic mRNA in cells can be improved, and the expression time can be prolonged.

The invention relates to the field and particularly relates to an expression vector for an animal cell. The expression vector sequentially contains a promoter with strong expression inductivity, a multiple cloning site for gene integration, a ribosome entry site sequence, a dihydrofolate reductase gene, a SV40 early stage promoter gene without an enhanser, a neo gene and an anti-drug gene from upstream. The invention further provides a preparation method of the expression carrier, a recombinant carrier and application. According to the expression carrier provided by the invention, expression of the neo gene is weakened by using the promoter without the promoter; a dhfr gene as a co-amplification gene is located at the downstream of the ribosome entry site IRES, so that the translation efficiency of the dhfr gene is reduced; the expression quantity of a target protein is increased by combining G418 pressurized screening and pressure screening of the dihydrofolate reductase gene.

The invention provides a leader sequence for improving non-cap-dependent translation efficiency and application of the leader sequence, and belongs to the technical field of genetic engineering. The leader sequence is as shown in SEQ ID NO. 1. According to the present invention, with the leader sequence, the expression quantity of the target gene mediated by the non-cap-dependent internal ribosome entry site (IRES) element in the transient mammalian cell expression system can be effectively improved;

The invention discloses an RNA (Ribonucleic Acid) composition for inducing neuronal differentiation. The RNA composition comprises a modified mRNA (Messenger Ribonucleic Acid) and a MicroRNA composition, the invention also provides a method for inducing differentiation of cortical glutamic acid energetic neurons by using the RNA composition, and pluripotent stem cells can be induced to be differentiated into neurons within seven days through transient overexpression of the mRNA and MicroRNA composition; the invention also provides an application of the RNA composition, and the RNA composition is used for inducing cortical glutamic acid neurons to differentiate. The transfection process and the mRNA delivery system are optimized, so that the translation efficiency of transfected mRNA and the expression quantity of mRNA in cells are remarkably improved. The method is suitable for inducing the differentiation of the cortical glutamic acid neurons, and the differentiated cortical glutamic acid neurons can be further applied to the construction of pathological models of human nerve-related diseases.