Invitrogen™ CorrectASE™ Enzyme

Catalog No.	Quantity
A14973	200 reactions

Catalog No. A14973 Supplier Invitrogen™ Supplier No. A14973

Description

CorrectASE™ enzyme removes mismatches caused by oligonuceotide synthesis errors, leading to a 3–10 fold reduction in mutations in your synthetic genes or fragments.

CorrectASE™ enzyme removes mismatches caused by oligonuceotide synthesis errors, leading to a 3–10 fold reduction in mutations in your synthetic genes or fragments.

By introducing CorrectASE™ enzyme into your do-it-yourself gene syntheis workflow, you can:

• Reduce the number of mutations in your synthetic gene or fragment
• Reduce your labor time by screening only 2–4 clones instead of 10-16 clones per synthetic construct
• Reduce your costs by sequencing only 2–4 clones instead of 10–16 synthetic genes

Prevent Unwanted Mutations

Commercially available synthetic oligonuceotides have a high error rate during synthesis, ranging from one per 300–1000 bases, depending on the source. These errors cause frameshift (deletion and insertion) and mismatch mutations during gene synthesis. Incubation with CorrectASE™ enzyme removes both type of mutations.

The incubation step with CorrectASE™ enzyme is introduced after the initial PCR assembly of oligonucleotides. The PCR product is denatured and reannealed so that any mutations will be unmatched. CorrectASE™ enzyme binds to the resulting mismatches and nicks both DNA strands 3' of the error. The 3' to 5' exonuclease activity of the enzyme removes the errors. A final PCR with a proofreading polymerase then assembles the corrected fragments, thus increasing the likelihood of isolating clones with the correct sequence. Depending upon the incoming oligonuceotide quality, only 2–4 clones need to be screened, compared to 10–16 clones in a workflow that does not include the correction step. Including CorrectASE™ enzyme in your gene synthesis workflow decreases labor time and sequencing costs.

Specifications

• Purity: >95% by SDS-PAGE
• Content And Storage: • CorrectASE, 4 tubes (50 reactions each)
  • 10X CorrectASE Reaction Buffer, 1 tube
• Exonuclease Activity: 3'–5'
• Enzyme: CorrectASE™
• Compatible Buffer: Reaction Buffer
• Quantity: 200 reactions
• Product Type: CorrectASE™ Enzyme

Frequently Asked Questions (FAQs)

What does CorrectASE enzyme do?

The CorrectASE enzyme has the ability to remove mismatches caused by oligonucleotide synthesis errors and is typically used in a do-it-yourself gene synthesis workflow to aid in decreasing mutations in your synthetic genes/fragements.

I'm confused with the primer concentration to use for the Do It Yourself Gene Synthesis Kit. Can you please clarify?

The protocol states that oligonucleotide stocks should be prepared at a final concentration of 100 µM in 1X TE buffer. The next line indicates the addition of 5 µL of each 10 µM primer together. According to R&D, the manual was written this way because our R&D typically brings up the lyophilized oligos to a 100 µM stock concentration (due to the volume of the tube). You do, however, want to use a 0.15 µM pool. Therefore, you can either dilute the stock to 10 µM or dilute the primer pool 1:1

I am not getting a PCR product using your GeneArt Do-It-Yourself Gene Synthesis Kit. What should I do?

Overdigestion with CorrectASE enzyme can lead to degradation of the DNA template.

Ensure that the reaction does not go longer than 60 mins. Also, ensure that the reaction is kept on ice until the PCR step or else the reaction will be prone to overdigestion by the CorrectASE enzyme.

I don't think my sequence is accurate. What should I do?

Please send an email to geneartsupport@lifetech.com explaining what is wrong. We will also need the project ID and construct ID, which we will forward to our QC department for further investigation.

After GeneArt Gene Synthesis, I cannot cut out my gene of interest using Xba1. Why is this?

We have a variety of strains that are used in production, such as Top10 or DH5α. Routinely, we grow them in dam+ strains. Therefore, you may be seeing inhibition because Xba1 is sensitive to dam methylation

For Research Use Only. Not for use in diagnostic procedures.