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View additional product information for GeneArt™ Strings™ DNA Libraries randomized bp - FAQs (833007DE)
11 product FAQs found
A GeneArt Strings DNA Library is a linear DNA fragment with the following parameters:
- 200 bp to 2000 bp in length
- Up to 3 randomized regions of up to 30 bp each
- The randomized regions can contain all IUPAC defined ambiguous nucleotides (N, K, S, B, etc.); U is not optional
- Randomized regions need to be at least 30 bp from either end of the fragment and at least 30 bp from each other
GeneArt Strings DNA Libraries are not available with TRIM technology or custom nucleotide mixes. Please inquire about our other library products if you require this type of randomization.
For GeneArt mutagenesis services, we will need the DNA or amino acid sequence of the gene you would like to mutagenize, the host organism you plan to use (this is important for gene optimization if you choose to include it with your request), the restriction sites you need at the 5'/3' ends and/or to avoid internally, and whether or not you want any other added motifs (e.g., Kozak sequence, stop codons, etc.).
Here are some examples:
- Increase or adjust promoter strength or specificity
- Enhance or modulate protein stability
- Modify or combine enzyme properties
- Increase binding affinities of receptors, ligands, and antibodies
- Optimize or alter signal peptide efficiencies
- Destroy protein function while retaining immunogenicity
- Combine and select natural polymorphisms
- Increase protein half life
- Adjust thermal stability
No material is necessary. For library creation, all we need is the sequence file, submitted electronically, and information about the position and nature of the sites you want to randomize. We can provide a quote for your project through our online ordering system (https://www.thermofisher.com/order/geneartgenes/projectmgmt). If needed, you can then relate detailed information about your library request to our production scientists prior to starting the project.
Yes, GeneArt Combinatorial Libraries can be randomized using any nucleotide mix you require, down to single digit percentages of given nucleotides (“dirty bottle” approach).
If you require a degenerate library with an unusual design please send an email to geneartsupport@lifetech.com. We will consider any project and in the vast majority of cases will find a solution to fulfill your requirements.
Combinatorial Libraries (up to 1,011 variants): Simultaneous randomization of multiple codons, TRIM technology optional
Site-Saturation Mutagenesis (up to 20 variants): Randomization of a single codon with every possible non–wild type variant
Sequential Permutation Libraries (# of codons x 20 variants): Successive site-saturation mutagenesis
Controlled Randomization (up to 1,011 variants): Unbiased random substitutions with defined frequency
Cloned cDNA of all known human SH3 domains
Truncation Libraries: Customer-defined truncations without out-of-frame mutations.
GeneArt Strings DNA Libraries: linear DNA fragments that can contain up to three regions of randomized nucleotides
There are a number of reasons:
- Because you already know that certain amino acid substitutions disturb the function of your protein (e.g., cysteines in complementarity determining regions (CDRs)).
- Because the number of possible variants of a protein is astronomically high, exceeding the capacity of even the highest-throughput screening capabilities by many orders of magnitude. The fewer useless mutations, such as those occurring in less important regions of the protein or that cause frame shifts or stop codons, the better your chances of finding a variant that results in the desired phenotype.
- Some screening assays are cost and labor intensive; thus, screening fewer clones saves time and money.
Conventional protocols for degenerated library creation (e.g., error-prone PCR) incorporate many unwanted mutations. Moreover, methods like DNA shuffling cannot typically cause recombination of directly adjacent mutations. Synthetic combinatorial libraries, on the other hand, limit the introduction of mutations to defined regions at the precise frequencies requested. In addition, adjacent mutations will be recombined (shuffled) independent of their proximity.
The answer depends on the specifics of your project. In general, it is advantageous to keep the diversity of a library as low as possible, targeting only the regions of a gene/protein that are likely to be functionally important. The following information can help determine this: crystal structure, conserved motifs, presence of homologs, etc.
We recommend this site: http://www.chick.manchester.ac.uk/SiteSeer/IUPAC_codes.htmL