Invitrogen GeneArt Strings DNA Fragments

The percentage of clones with correct length as identified by colony PCR (cloning efficiency) and the percent of sequence-correct clones (fidelity) based on the number of full-length clones.

Eight Strings DNA Fragments of different lengths up to 2,800 bp with suitable sequence ends (AarI recognition sequence and 6 bp stuffer nucleotides) were used for direct assembly with the GeneArt Type IIs Assembly Kit, Aar I. The resulting four genes were 5,056 bp, 5,061 bp, 5,267 bp, and 5,448 bp long. After ligation and transformation, bacteria were plated and incubated overnight at 37°C. Sixteen colonies were picked, and colony PCR was performed to identify full-length clones. For all four assembled genes, eight colonies were sequenced to determine the number of sequence-correct clones.

Ten Strings DNA Fragments up to 3,000 bp were cloned using the GeneArt Seamless Cloning and Assembly Kit. After ligation and transformation, bacteria were plated and incubated overnight at 37°C. Eight (<1 kb) or 16 (>1 kb) colonies were picked, and colony PCR was performed to identify full-length clones; four to eight full-length clones were sequenced to determine the number of sequence-correct clones.

While Strings DNA Fragments are offered in lengths up to 3,000 bp, GeneArt Seamless assembly technology can be used to directly assemble two Strings DNA Fragments up to 1 kb without pre-cloning of the subfragments.

If you want to assemble more Strings DNA Fragments or longer Strings DNA Fragments using seamless assembly technology, pre-cloning of the subfragments is recommended to limit post-assembly screening efforts necessary for finding correct clones. Alternatively, you can use GeneArt Type IIs assembly technology.

Five Strings DNA Fragments up to 1,000 bp with attB sites were cloned into pDONR 221. After ligation and transformation, bacteria were plated and incubated overnight at 37°C. Eight colonies were picked, and colony PCR was performed to identify full-length clones; four to seven full-length clones were sequenced to determine the number of sequence-correct clones.

Strings DNA products can be cloned using any available cloning method, provided the appropriate ends have been included in the design. End sequences can be added prior to uploading the sequence into the GeneArt Services Dashboard or they can be added after import.

Strings DNA Fragments are PCR amplicons of assembled oligonucleotides, ready for screening to identify the correct clone. Strings DNA Fragments are bulk sequenced as part of the quality control process to help ensure that your desired sequence is present in the fragment pool.

To identify a correct clone >90% of the time, we recommend using the following screening guidelines.

GeneArt Strings DNA Fragments are custom-made, uncloned, double-stranded linear DNA fragments, available in lengths from 200 base pairs (bp) to 3,000, assembled from synthetic oligonucleotides using the same process developed for GeneArt high-quality gene synthesis. Strings Fragments can be used to quickly clone your gene of interest. At least 200 ng of Strings DNA Fragments are produced within 3-5 (up to 1,000 bp) or 8 (from 1,000 to 3,000 bp) business days. GeneArt Strings DNA Fragments can be designed, sequence optimized, and ordered online and are a cost-effective and smart alternative to clone your expression plasmid.

While we used to offer Strings DNA Fragments between 100 – 199 bp, this is unfortunately no longer possible.

Strings Fragments are amplicons (PCR amplificates) of assembled oligonucleotides; an intermediate product of the gene synthesis production process. This process results in a pool of fragments, and cloning and screening need to be carried out to identify the correct clone. To ensure that correct fragments are present in the pool, Strings Fragments are bulk sequence-controlled before shipment.

Strings DNA Fragments are shipped dried, ready for resuspension and cloning. Before using, we recommend that you spin down the tube contents, add some water to the bottom of the tube to get the desired concentration, and let it incubate for at least 1 hour at room temperature (alternatively incubate at 4°C overnight). Subsequently, resuspend carefully and use immediately. For longer storage, resuspended Strings DNA Fragments should be dispensed into aliquots and frozen at –20°C. Please avoid freeze-thaw cycles.

No. Strings DNA Fragments from 200 to 1,000 bp are produced in 5 business days and Strings DNA Fragments from 1,000 to 3,000 bp are produced in 8 business days. Depending on the nature of the sequence, production time can vary.

Yes. Since you need to perform the cloning to obtain your final gene, Strings DNA Fragments prices are always below those for gene synthesis.

Yes. We recommend using the GeneArt web order portal for ordering, where you can use the free gene editing and optimization functions to adjust the sequence to meet your needs. After editing and optimization, please check again that the final sequences of the fragment(s) are exactly as needed for further processing in your lab (e.g., potential homologous overlaps of the fragments or restriction sites and buffer bases needed for cloning).

No, subcloning and assembly services are not available for Strings DNA Fragments, as they are designed to offer you the fastest and most affordable way to get access to your genes. If you do not want to clone your gene yourself, we recommend that you order full gene synthesis service.

Yes, it is possible to directly assemble 2 Strings Fragments without pre-cloning. Thermo Fisher Scientific offers several technologies for seamless assembly, e.g., GeneArt Type IIs Assembly or GeneArt Seamless Cloning and Assembly. Please refer to the application examples section on this page for more information. Depending on the sequence length and the number of subfragments you want to assemble, the screening effort to find a correct clone can be high. We therefore generally recommend pre-cloning the subfragments to limit the screening effort required to find a correct clone after assembly.

No. Strings DNA Fragments are limited to 200 to 3,000 bp. If your sequence is longer, you need to order it as gene synthesis or assemble your gene from two or more Strings DNA Fragments. In addition, the Strings DNA Fragments manufacturing process is streamlined to very quickly and cost-effectively provide you with your gene product. If your sequence is complex, you may receive a message that it cannot be produced as a Strings DNA Fragment due to high complexity. In that case, we recommend modifying your sequence to match the production criteria (given below) or ordering your gene as gene synthesis. In many cases, optimizing your sequence using the portal function enables manufacturing with the Strings Fragments process. If it is still not able to be produced, you can manually edit the sequence to enable production.

Important production criteria are:

• GC content between 20% to 80% (in peaks, not overall content)
• No long secondary structures or sequence repetitions
• No A/T stretches >25 bp nor G/C stretches >20 bp