GeneArt® Strings™ DNA Fragments


GeneArt® Type IIs Assembly

GeneArt® Strings™ DNA Fragments


Q. In basic terminology, what are Strings?

A. GeneArt® Strings™ DNA Fragments are custom-made, linear, double-stranded DNA fragments up to 3 kb in length, ready for cloning in your lab. Please refer to www.lifetechnologies.com/strings for full details.


Q. What is the price per base?

A. GeneArt® Strings™ DNA Fragments are priced in length categories, not per base pair like gene synthesis. The average price is around $0.2 per base pair, so it’s always 20–30% below gene synthesis price.

Product Price
Strings™ DNA Fragments 100–600 bp $99
Strings™ DNA Fragments 601–750 bp $129
Strings™ DNA Fragments 751–1,000 bp $149
Strings™ DNA Fragments 1,001–1,250 bp $219
Strings™ DNA Fragments 1,251–1,500 bp $259
Strings™ DNA Fragments 1,501–1,750 bp $299
Strings™ DNA Fragments 1,751–2,000 bp $349
Strings™ DNA Fragments 2,001–2,250 bp $399
Strings™ DNA Fragments 2,251–2,500 bp $449
Strings™ DNA Fragments 2,501–2,750 bp $499
Strings™ DNA Fragments 2,751–3,000 bp $549


Q. Can you explain how the error correction is done, and what enzyme is used?

A. Error correction is done by an enzymatic step during PCR amplification of GeneArt® Strings™ DNA Fragments. We cannot disclose the enzyme we use due to proprietary information.


Q. When you compared traditional vs Strings™ Fragments, you mentioned the steps after amplification can be avoided. How do you sequence the fragment then? How do you confirm the sequences are correct?

A. GeneArt® Strings™ DNA 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 help ensure that correct fragments are present in the PCR product, GeneArt® Strings™ DNA Fragments are bulk sequence-controlled before shipment. GeneArt® Strings™ DNA Fragments are only sent to customers if we can verify that the customer desired sequence is present in the fragment pool.


Q. How do you deal with confidential sequences?

A. Confidentiality is maintained at our facility by strict adherence to customer confidentiality policies, ensuring all production is carried out in-house using the same procedures for GeneArt® gene synthesis.


Q. Codon usage optimization is a great service when it's needed, but it's possible to skip codon adaptation optimization and get exactly what we order as well, right?

A. Yes, you can simply paste your desired sequence into the portal editor (or Excel order sheet if you prefer to order outside the portal) and order without optimization or any sequence modification.


Q. Is the optimization process a genetic algorithm or does it output a consistent result for a given input?

A. If you do not change any parameter, the algorithm delivers the same result for a given input. However, we do constant improvements based on our research and literature data on our algorithm that may lead to different results if you optimize at different time points.


Q. Does the optimization function change the amino acid sequence of my gene?

A. No. The amino acid sequence is not changed by gene optimization.


Q. If I order two complementary strands, what is the recommended way of hybridizing them that give higher percent of correct double strand? Is it better to order to smaller (800 bp) oligos or longer (3 kb each) oligos when trying to clone them into a vector?

A. GeneArt® Strings™ DNA Fragments are double-stranded DNA molecules, it is not possible to order complementary single strands. We recommend ordering one larger sequence instead of several shorter ones for assembly, since this will be less work for you.


Q. On slide 29, what was the difference between 5448 (100% efficiency) and the other 2-Strings™ Fragment assemblies?

A. There was no specific difference, this just exemplifies the range of results you can have. 100% cloning efficiency just means in this case we found all colonies we screened by colony PCR to have the correct length.


Q. Do I need to cut the Strings™ Fragments if I want to do restriction enzyme cloning? And if I want to do TOPO® cloning, do I need to run PCR to get the overhangs?

A. Yes, GeneArt® Strings™ DNA Fragments are delivered as double-stranded blunt PCR fragments and you need to digest to get the desired overhangs or PCR amplify to get the A-overhangs for TA cloning. No need for any modification for TOPO® cloning, but here we recommend to add 5–7 nucleotides of stuffer sequence to protect the ends against naturally occurring truncations and to increase the chance of obtaining full-length clones.


Q. Is the fidelity rate very high?

A. The sequence correctness (or fidelity) is high, but depends on the length of the fragment. You will find one correct clone with high likelihood if you stick to the screening recommendations given on the webpage: sequence 2–4 full-length clones for sequences up to 1 kb, 3–5 full-length clones for sequences 1–2 kb, and 4–8 full-length clones for sequences 2–3 kb. Full-length clones can be identified by colony PCR on the picked clones.


GeneArt® Type IIs Assembly


Q.How many reactions can you do with one GeneArt® Type IIs Assembly kit?

A. 10 reactions.


Q. Do you plan on adding any more Type IIs enzymes in the future?

A. There are currently no plans to do so.


Q. What was the quality of the 8-fragment cloning?

A. With careful design, approximately 50–80% cloning efficiency can be achieved if pre-cloned fragments are used in the assembly reaction. The efficiency may be affected by multiple factors such as nature of sequence, DNA purity, etc.
See examples


Q. There are several different cloning kits on the market. What is the uniqueness of GeneArt® Type IIs assembly? Or, to what application is this kit aimed?

A. The GeneArt® Type IIs assembly kits have the advantage of seamlessly assembling multiple repetitive sequences and short fragments, unlike what is typically possible with homologous recombination–based assembly kits that are available commercially.


Q. What is the maximum length of PCR fragments that can be used for Type II assembly?

A. We recommend <2 kb mainly due to the robustness of polymerases on the market. Although we recommend using the GeneArt® Type IIs Assembly Kit for assembling up to 8 DNA fragments plus a vector, totaling up to 13 kb in length, you can use it to create constructs that are up to 20 kb in size; however, the cloning efficiency and the number of transformants will be lower.


Q. What would be the advantage of using Type IIs assembly over Gibson method?

A. The GeneArt® Type IIs Assembly Kits have the advantage of seamlessly assembling multiple repetitive sequences and short fragments. Gibson assembly, while also scarless, depends on the overlapping (homologous) ends of adjacent fragments, so repetitive sequences or fragments with unwanted internal homology may reduce efficiency.


Q. What are the size ranges you have tested for 5+ part assembly, and what efficiencies do you obtain?

A. Using fragments sized up to 2 kb each, we expect 50–80% cloning efficiency can be obtained with careful design.


Q. How does the efficiency compare to other advanced cloning kits like TOPO® TA Cloning® kits, etc?

A. TOPO® TA Cloning® kits are usually only for cloning one fragment into a vector, and usually leaves scars (unwanted sequences), while GeneArt® Type IIs assembly kits can seamlessly assemble multiple fragments.


Q. Can we get the .pptx file for this presentation?

A. View a recording of the webinar here.