Gene Design
Expression of a native gene in a heterologous host often involves sequence preference incompatibility, which may result in low or no expression. Eurogentec’s Gene-Optimizer™ Design system is an optimisation technology that can modify gene sequences to achieve the highest possible levels of productivity in any given expression system. The Gene-Optimizer™ algorithm takes account of a variety of critical factors involved in different stages of protein expression, such as codon adaptability, mRNA structure, GC content, repetitive sequences and various others cis-elements in transcription and translation.
Eliminating unfavorable and uneven GC content: The strength of the guanine-cytosine bond can lead to undesirable mRNA secondary structures.
Avoiding unfavorable mRNA secondary structures: Overly stable mRNA secondary structures, particularly at the 5' end of the transcript, have been implicated in reduced gene expression. The potential of a transcribed mRNA to adopt such a structure can be identified using free energy calculations.
Selecting a codon usage table: Eurogentec can design genes based on the codon usage tendencies of any requested organism. For proteins that are to be expressed in more than one host, Eurogentec can create hybrid codon usage tables.
Amino Acid | Codon | E | Y | D | H | | |
| Phe | TTT | 0.58 | 0.59 | 0.37 | 0.45 | | E: E.coli |
| | TTC | 0.42 | 0.41 | 0.63 | 0.55 | | Y: Yeast |
| Tyr | TAT | 0.59 | 0.56 | 0.37 | 0.43 | | D: Drosophila |
| | TAC | 0.41 | 0.44 | 0.63 | 0.57 | | H:Human |
Phenylalanine (Phe) and Tyrosine (Tyr) amino acids codon usage bias in 4 different organisms
Adding or removing restriction sites: The presence or absence of selected restriction sites is often important to facilitate subsequent gene manipulations such as swapping between vectors, exchanging protein domains, and adding or removing peptide tags or fusion partners.
Adding or removing specific constraints: Insertion or deletion of regulatory elements (polyadenylation signals, RNA methylation signals, selenocystein signals), adding or removing immuno-stimulatory or immuno-suppressive elements (for DNA vaccines) can be easily realised.
Creation of gene variants (mutant forms): Multiple variant forms (or variant library) can be designed for functional study and screening..
Applications of Custom Genes
Custom Genes technology is extremely versatile and applicable in many different kinds of applications, such as cDNA production, heterogenous protein expression, and the creation of gene variants and recombinant antibodies.
Gene Modifications: The presence or absence of selected restriction sites is often important to facilitate subsequent gene manipulations such as swapping between vectors, exchanging protein domains, and adding or removing peptide tags or fusion partners.
Gene Variants and SNPs: Most alternative splice methods are unwieldy and can have unpredictable results. Custom genes synthesis incorporates rigorous regulation and verification of alternative splice products.
RNA optimisation: Overly stable mRNA secondary structures, particularly at the 5’ end of the transcript, have been implicated in reduced gene expression. Avoid unfavourable RNA secondary structure and increase your protein production.
Codon optimisation: Not all host expression systems share the same codon bias. The expression of gene of interest increases dramatically when the codon frequency of that gene is matched to that of the host system. For proteins that are to be expressed in more than one host, Eurogentec can create hybrid codon usage tables.
cDNA Design: Eurogentec’s gene synthesis technology is developed for the design of high-quality cDNA for cDNA libraries and other purposes.
Microarray-Ready cDNA: Reverse transcription bias and other problems can alter the relative amounts of cDNA measured by microarray assays.
Recombinant Antibodies: Human-mouse chimeric antibodies retain the specificity of the original murine variable region but assume the effector function of the human constant region, combining the high specificity of hybridoma antibodies with the mutability and reliable replication of plasmids.
DNA Vaccines and Vectors: The production of an effective DNA vaccine involves not only the sequence itself but also the target, delivery system, expression, regulation, and inhibition of the gene therapy product.
Predicted Gene: The primary reason to generate predicted gene structure databases is to provide positional cloners, gene hunters and other products along with the gene candidates observed in finished and unfinished genomic sequences. Eurogentec specializes in custom gene synthesis and predicted gene synthesis based on advances in the understanding of gene variant function and synthesis, the biology of gene synthesis and new DNA manipulation and mutagenesis techniques.
Custom Genes Synthesis
 | You create your own Custom Gene(s) (up to 50 kb) with your sequence(s) and the restriction sites to facilitate its integration in your vector even its expression by the chosen host. |
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 | Please, send the Excel file named custom genes.xls (Eurogentec website) filled with all the details associated with your Custom Gene(s) to custom.gene@eurogentec.com |
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 | On receipt of your complete order (PO number, invoice and delivery addresses...), Eurogentec will synthesise your Custom Gene(s) with the following steps: synthesizing oligonucleotides, obtaining of full-length sequence(s), correcting possible mutations. |
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 | Your Custom Genes will be cloned in pUC57 vector by default, to allow its amplification. The cloning is also possible in other vectors, commercial one or vector built for your experiments (additional charge). |
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 | Your Custom Gene(s) is Quality Controlled by several methods to ensure: - Size of the inserted fragment (digestion and PCR);
- 100 % sequence of your Custom Gene(s) (sequencing);
- Reading frame (if requested);
- Flanking regions of the chosen vector (sequencing);
- DNA quality and quantity (spectrometry).
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Delivery Specifications
After synthesis you will receive 4 μg of lyophilised plasmid DNA carrying your Custom Gene. Each of your Custom Gene will be accompanied by various documentations:
- A printout of the sequence alignment after sequencing;
- A plasmid map to locate the flanking regions and restrictions sites;
- General information and instructions to directly use your Custom Gene(s);
- Quality Assurance Certificate.