Documenting a construct so the next person can use it

The next person should be able to open one sheet and see the frame, the antibiotic, the strain, the sequence file, and the freezer slot without asking you. You are writing a minimal construct record, and the words on it have to mean one thing each. This glossary defines those words by building the sheet. It is a research note, not a laboratory information system and not a claim that the plasmid is correct merely because the boxes are filled.

How the colony was earned is in plasmid cloning from insert to colony. Where a cold box should live is in storing biological samples from fridge to freezer.

The sheet is a set of definitions

A map file is the annotated sequence the next person can open: bases, features, and a version. A GenBank-style record is the usual class. NCBI GenBank is where a public parent accession may live. Your construct may only exist as a local file. Name that file on the sheet. A slide with a coloured arrow is not a map file. The frame has to be visible on it. Mark the start that is actually translated, any tag fusion, and the junction bases. "In frame" without those coordinates means the next person must guess which ATG you trusted.

Primers are the oligonucleotides that built the insert or that read it, written as names plus sequences, with the coordinate they bind on this map. A primer name from another project is a trap. If restriction ligation used a pair of sites, name the enzymes beside the primers that introduced them. If the orientation check was a primer that only amplifies one direction, say so. The next person should not have to reconstruct your PCR from a freezer box of unlabelled tubes.

The antibiotic is the resistance marker on the backbone and the drug you actually poured. Write the name you used. A backbone that carries two markers needs the one this stock was selected on. The concentration belongs in the SOP you cite, not as a remembered number from a different plasmid. Antibiotic on the sheet is what stops someone reviving the stock on the wrong plate and concluding the plasmid died.

The strain is the host genotype that changes the result. Dam and Dcm status, a recombination deficiency, and whether the cells tolerate a counterselection cassette are the kinds of facts that belong here. "E. coli" is not a strain. The next transformation has to be able to copy the strain line onto a competent-cell request.

The date is the day this archive was made from the colony you are willing to defend. A date on an old gel and a date on a glycerol tube that was restreaked later are two dates. Use the date of the material in the slot. If you revived and refroze, add a line. Do not leave the original day pretending to describe a later scrape.

The sequence file is the chromatogram or the assembled read, named so it can be found, and the statement of how it compared with the map. "Sequenced" is not a file. If one read covered the insert, say which primer and which coordinates. If the plasmid is longer than that read, the unread span goes to the next field.

The glycerol location is the freezer, the box, and the slot of the living stock, and it should point at a plasmid DNA tube as well if you keep one. The storage article decides the temperature class. This field decides which hole. A location that says "my box" dies when you leave.

What was not confirmed is a required line, not an apology. It lists the checks you did not do: an unread junction, orientation not tested, a single colony sequenced, no expression trial, no digest after the last revival. The next person can then choose to repeat a check or to limit their claim. An empty line is how a partial record impersonates a finished one.

Fill it when the colony is still the one you read

Start the sheet when the sequence is compared with the map, not months later from memory. Copy the filename, the strain on the competent-cell tube, and the antibiotic on the plate you picked from. Freeze the glycerol stock and put the slot on the sheet the same day. If a field is unknown, write unknown. Do not invent a strain from the plasmid's reputation.

Branch if the read disagrees with the map. The sheet can still exist, under a name that states the disagreement, with the sequence file attached. Do not file it under the intended construct's name. The next person will use the name you chose.

Branch if you have a map and no living stock. Record the plasmid DNA tube and write that no glycerol location exists. They can transform later. A blank location looks like a lost box, which sends people searching a freezer for a stock you never made.

Branch if orientation was not tested. That fact goes under what was not confirmed, and the map arrow stays unmarked or marked as unchecked. Restriction ligation of a blunt insert is the classic case. The next person can design the missing check instead of expressing the backwards gene for a month.

Failures the sheet prevents

Guessing the frame from a gene name is how a tag fusion is translated out of frame. The map field exists so the junction bases are visible without you. Primers stored only as a spreadsheet on one laptop fail the same way when that laptop is gone. Write the sequences into the sheet or into the map file the sheet names.

Two stocks with one name and no dates are how a mutant revival replaces the sequenced colony. The date and the sequence filename are the pair that stops it. The glycerol location without a date is a hole in a box, not a genotype.

A sheet that omits what was not confirmed trains the next person to over-claim. They will write "fully sequenced" in their own talk because your sheet looked complete. The negative line is the courtesy.

Safety and the record

If the insert has a hazard class, the sheet points at the institutional decision. It does not grant one. The WHO Laboratory biosafety manual, 4th edition is background. Antibiotic-resistant stocks are waste when discarded, and they are labelled material when kept. A nickname tube in a shared freezer is how the wrong plasmid is grown by someone who shares the room.

Boxes move

In a shared cold room, racks are relabelled and boxes are consolidated. The location line has to be updated by the person who moved the box, the same day. A perfect map with a stale slot is a plasmid nobody can use. Write the slot in the form freezer, box, position. When you leave, the sheet should still find the tube.

What you attach to a later enquiry

The sheet is the specification. Enzymes for a rebuild sit in the molecular biology catalogue. A junction that was never read is a sequencing request: use the Sanger sequencing enquiry reference and attach the map plus the "not confirmed" line. If the next person must have the plasmid respecified from the file rather than from a doubtful stock, the custom gene synthesis enquiry reference is only a prompt for that file. Send the sheet with the quote request. Ask whether a quotation is possible. A gene name is not the attachment.