Construct system and uses therefor

Abstract

The present invention discloses construct systems and methods for comparing different iso-accepting codons according to their preference for translating RNA transcripts into proteins in cell or tissues of interest or for producing a selected phenotype in an organism of interest or part thereof. The codon preference comparisons thus obtained are particularly useful for modifying the translational efficiency of protein-encoding polynucleotides in cells or tissues of interest or for modulating the quality of a selected phenotype conferred by a phenotype-associated polypeptide upon an organism of interest or part thereof.

Description

FIELD OF THE INVENTION[0001]The present invention relates generally to gene expression. More particularly, the present invention relates to construct systems and methods for comparing different iso-accepting codons according to their preference for translating RNA transcripts into proteins in cell or tissues of interest or for producing a selected phenotype in an organism of interest or part thereof. The codon preference comparisons thus obtained are particularly useful for modifying the translational efficiency of protein-encoding polynucleotides in cells or tissues of interest or for modulating the quality of a selected phenotype conferred by a phenotype-associated polypeptide upon an organism of interest or part thereof.BACKGROUND OF THE INVENTION[0002]The expression of foreign heterologous genes in transformed cells is now commonplace. A large number of mammalian genes, including, for example, murine and human genes, have been successfully expressed in various host cells, includ...

AI Technical Summary

Benefits of technology

"The present invention provides a method for improving the sensitivity of determining the translational efficiency or phenotypic preference of different synonymous codons by using a construct system that employs different reporter polynucleotides that code for the same amino acid sequence. The system includes a plurality of synthetic constructs that each contain a reporter polynucleotide and a unique interrogating codon that codes for a specific amino acid. The interrogating codons differ from one another in their choice of position in the amino acid sequence. The system can also include additional synthetic constructs to interrogate the translational efficiency or phenotypic preference of additional interrogating codons. The coding sequence of individual reporter polynucleotides can be designed to confer a phenotype upon a cell or tissue in which it is expressed."

Problems solved by technology

Nevertheless, despite the burgeoning knowledge of expression systems and recombinant DNA technology, significant obstacles remain when one attempts to express a foreign or synthetic gene in a selected host cell.

This lower than expected translation efficiency is often due to the protein coding regions of the gene having a codon usage pattern that does not resemble those of highly expressed genes in the host cell.

It is widely known in this regard that translation of “rare codons”, for which the corresponding iso-tRNA is in low abundance relative to other iso-tRNAs, may cause a ribosome to pause during translation which can lead to a failure to complete a nascent polypeptide chain and an uncoupling of transcription and translation.

Thus, the expression of an exogenous gene may be impeded severely if a particular host cell of an organism or the organism itself has a low abundance of iso-tRNAs corresponding to one or more codons of the exogenous gene.

In other words, it was discovered that different cells of an organism can exhibit different translational efficiencies for the same codon and that it was not possible to predict which codons would be preferred, less preferred or non preferred in a selected cell type.

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