What to send for a gene synthesis enquiry

A gene-synthesis enquiry can be checked when six things are in the request: the sequence, the vector, the host, the sites to preserve or avoid, the expression constraints, and the biosafety of the DNA. You are troubleshooting an incomplete request, the way you would troubleshoot an empty plate: find the missing fact, and do not invent it. A quotation built on a gene nickname cannot be checked against the molecule you will clone.

How a laboratory writes a sourcing note in general is in how to write a laboratory sourcing enquiry. How the fragment becomes a verified colony is in plasmid cloning from insert to colony. The custom gene synthesis enquiry reference is a place to point this specification. It is an enquiry reference only.

What a checkable request is made of

The sequence is the full bases you want, in a file with a version. Include the translation if an open reading frame must stay in frame. A sentence such as "the human version" is not a sequence. Look up the accession you mean in NCBI GenBank and attach that record's identifier plus any edits. If you already blasted the file against the wrong organism, say what NCBI BLAST returned so the identity is explicit.

The vector is the backbone the fragment should end in, or a clear statement that you want linear DNA and will clone it yourself. Name the marker, the copy number if it matters, and the map. "A standard expression vector" is not a vector. Two people will picture two plasmids.

The host is the species and the strain that will carry or express the construct. Codon choices have no target without it. A cloning strain and an expression strain are different lines on the form. Write both if both will see the plasmid.

Sites to preserve or avoid are a list with reasons. A restriction site you will use next month has to survive. A site inside a coding sequence that collides with that plan has to be removed without changing the protein, or you have to accept the amino-acid change. Silence is how those two instructions contradict each other inside one quotation.

Expression constraints are the promoter context, tags, fusion frame, secretion signal, and any codon preference you actually need. "Optimise it" without a host and without a list of residues you must not change is not a constraint. It is a request for someone else to guess.

Biosafety of the sequence is a plain description of what the product is, the source organism, and whether it is a toxin, a virulence factor, or otherwise under a rule your institution already applies. That disclosure is what lets screening and local approval happen in the open. This article is not a way to shape a sequence so a check misses it. If the identity is sensitive, the enquiry stops until the right committee has seen the real bases. The WHO Laboratory biosafety manual, 4th edition is background for how laboratories think about biological risk. It is not an approval for your insert.

Walk the draft and stop at the first hole

Write the six fields in that order before you ask for a quotation. Then read the draft as if you had to clone from it with no further email.

If the sequence is missing, stop. Nothing else can be aligned to a file, and a quotation has nothing to be checked against. Do not send a gene symbol and a request to use the common one.

If the sequence is present and the vector is missing, stop before codon work. The ends of the fragment depend on whether it will be ligated, assembled, or left linear. Add the map or state that the vector is undecided and list the features it must have.

If the host is missing, stop before any codon edit. A change that suits one expression species can be pointless in another. Name the strain that must tolerate the plasmid, especially if a counterselection cassette or an unstable insert is involved.

If sites are missing, look at the cloning method you wrote under the check. Restriction ligation needs the sites you will cut. Gibson assembly needs overlaps that are not repeated elsewhere in the construct. Add the list, or write "no site constraint" so the absence is a decision.

If expression constraints contradict the sites, stop. An example is a site you asked to preserve that sits in a codon you also asked to change. Resolve it in the file. A quotation cannot contain both instructions.

If biosafety is blank, stop. Do not fill it with "non-hazardous" as a habit. State the product. If you do not know whether the product is regulated, say that uncertainty and name the institution that will decide. An evasive description is a failed request.

Failures that survive into the clone

A complete-looking request can still be wrong. The sequence file and the translation disagree. Trust neither until you reconcile them. Colony screening will then confirm a molecule that matches the wrong instruction.

The vector map is from an old version whose multiple cloning site has changed. The sites you asked to preserve are not in the plasmid you will receive. Attach the map you mean, not a name alone.

The host on the form is the expression species, and the cloning strain is a different genotype that cannot maintain the backbone. Write both. Otherwise the first transformation fails for a reason the quotation never contained.

You asked for a short overlap for Gibson assembly and also for a restriction site in the same ten bases. Those ends fight. Pick the joining method and let the enquiry describe those ends only.

An incomplete biosafety line that someone "filled in later" with a milder description than the file is a different molecule from the one the committee would have seen. Keep the description and the bases the same document.

Research use and institutional rules

This page does not approve synthesis, cloning, or expression. Diagnostic, clinical, and forensic claims are out of scope. Containment, transport, and whether a sequence may be made at all are institutional and legal questions. Ask them with the real sequence attached. Waste and recombinant plates, once any DNA exists, follow the same local rules as the rest of your cloning.

A specification a second site can read

The practical test is whether another laboratory could clone from the request without calling you. That is the same habit as any sourcing note: name the method class, the acceptance check, and the identity of the material. A sequence pasted into a chat with no file version cannot be matched to a later quotation. Attach the file. Refer to it by name and date. Use the quote request for the ask, and point at the custom gene synthesis enquiry reference only as the subject line for that kind of specification.

Enzymes you will use after the fragment exists, ligase or an assembly mix, can be listed from the molecular biology catalogue in the same note so the ends and the enzyme class agree. Ask whether a quotation is possible. Send the six fields. A gene name and a deadline are not a sixth field.