The High Accuracy of ENFINIA™ Linear DNA
Stop Screening Clones. Start Using DNA.
The Error Rate of DNA Synthesis Matters
DNA accuracy directly impacts the speed and reliability of experimental progress. This is because synthesis errors dilute the population of perfect DNA molecules, such that the product contains desired perfect sequences and also undesired imperfect ones that can skew the results of experiments. DNA length can also impact the percentage of perfect molecules, as longer DNA is more likely to contain at least one error than shorter DNA.
While conventional DNA fragments are inexpensive and fast to produce, most non-clonal gene fragments on the market are produced with a 1:5000 error rate or higher. For a 3 kb gene, only about half of its molecules would actually be sequence-perfect.
When fragments contain sequence errors, researchers must spend additional time identifying a correct construct. This typically involves cloning the fragment into plasmids, transforming bacteria, growing colonies, and sequencing multiple clones to find an error-free sequence. Only then can the correct clone be amplified and purified for downstream use. Longer sequences with more errors require more cloning to identify a perfect copy to use in downstream experiments. These additional steps can add a week or more to an experimental workflow.
Lower synthesis error rates increase the likelihood that a DNA molecule is already sequence-perfect, reducing the need for cloning and colony screening and enabling researchers to move more quickly from sequence design to experimental testing.
Conventional DNA fragments require cloning. Due to higher error rates, conventional DNA fragments require bacterial cloning to identify the exact sequence.
High-Accuracy Linear DNA: Skip Cloning and Save Time
Historically, to avoid cloning DNA fragments and screening colonies in the lab, researchers had to outsource clonal gene synthesis, whereby vendors clone DNA into plasmids for you. However, this option takes longer and costs more.
ENFINIA Linear DNA offers an alternative—combining the speed and cost of fragments with accuracy approaching clonal DNA. With a median error rate of 1 error per 70,000 bp, ENFINIA Linear DNA achieves 99.999% per-base accuracy. For a standard complexity 3 kb construct, approximately 96% of molecules are sequence-perfect, allowing constructs to be used immediately upon arrival—without the need for cloning or colony screening.
- 1 error / 70,000 bp
- ~96% sequence-perfect molecules
- NGS-verified constructs
- Delivered in as few as 6 days
- Low cost
The exceptionally low error rate of ENFINIA DNA is attainable because of Elegen’s proprietary cell-free cloning process, in which DNA molecules are assembled enzymatically from oligonucleotides in a controlled, cell-free environment optimized for sequence fidelity. The high accuracy of constructs is maintained for sequences up to 7 kb and across a broad range of complexities. In addition, every construct is NGS-verified for sequence accuracy and purity before shipment.
Since ENFINIA Linear DNA can be used immediately without cloning, it is the fastest and most cost-effective DNA for researchers (see graphic).
ENFINIA Linear DNA vs. fragments and plasmids. ENFINIA Linear DNA combines the speed and cost of fragments with the accuracy of plasmids. Delivered in as few as 6 days, NGS-verified, and ready to use without cloning for most applications.
High Accuracy Synthesis without Cloning in Cells
DNA synthesis error rates are often lower than the intrinsic error rates of most sequencing platforms. As a result, true synthesis errors can be masked by sequencing noise, making it difficult to measure DNA synthesis error rates directly using standard sequencing methods.
To overcome this limitation, Elegen’s error quantification method uses Unique Molecular Identifiers (UMIs) and long-read sequencing to distinguish true synthesis errors from PCR or sequencing artifacts. This approach enables the distinction of true synthesis errors from sequencing noise with high confidence, enabling accurate quantification of ENFINIA Linear DNA error rates.
A: Starting population
A population of identical DNA molecules may contain rare errors introduced during synthesis.
B. Molecular tagging
Before PCR amplification, each original DNA molecule is tagged with a UMI.
C. Amplification
PCR amplifies the DNA along with its UMI, creating a family of reads derived from the same original molecule.
D. Sequencing and analysis
After long-read sequencing, reads sharing the same UMI are grouped together. If a mutation appears in all reads within a UMI group, it originated in the original molecule and represents a true synthesis error. If it appears in only a subset of reads, it is likely a PCR or sequencing artifact and is filtered out computationally.
Detecting True Synthesis Errors
The parental DNA molecule with the “light blue” UMI has a true synthesis error, as shown in B & C (indicated by the small red rectangle). Because the parental molecule is amplified, the signal for this mutation is amplified such that nearly all the reads conflict with the reference sequence. Because the mutation is present in all reads, it can be attributed to a synthesis error rather than a sequencing error.
Elegen’s High-Quality DNA
Using this UMI-based method, Elegen measured synthesis accuracy across 45 customer sequences produced through the standard commercial workflow. These sequences included both standard sequences and high-complexity sequences (high GC content, local GC variation, repeats).
Across all samples, ENFINIA Linear DNA demonstrated a median error rate of 1 in 70,000 bp. This accuracy is more than an order of magnitude better than typical DNA fragments from major suppliers.
DNA You Can Use with Confidence
ENFINIA Linear DNA combines the speed and affordability of fragments with the accuracy of clonal DNA.
Powered by Elegen’s cell-free synthesis platform and verified by NGS, ENFINIA Linear DNA delivers long constructs that are highly accurate and ready to use upon arrival.
Delivered in as few as 6 business days and priced competitively, ENFINIA Linear DNA offers a robust alternative to conventional low-accuracy DNA fragments. It’s time to skip cloning and get straight to experiments.
Check out how ENFINIA Linear DNA can save you time.