The most expensive email a sequencing provider can send you is "your samples failed QC". You lose weeks, sometimes pay for re-QC or new library prep, and sometimes lose samples you can never collect again. Most failures come from a few avoidable mistakes. Here is how to catch them before you ship.
01Measure the amount the way the provider will
A NanoDrop measures everything that absorbs light at 260 nm: DNA, but also RNA, free nucleotides and some leftover extraction chemicals. That's why it often overestimates DNA. Providers re-measure with a fluorometer (Qubit or similar), which uses a dye that binds only the molecule you care about.
02Know the common QC targets
| Measure | DNA | RNA |
|---|---|---|
| A260/280 (protein) | about 1.8 to 2.0 | about 1.9 to 2.1 |
| A260/230 (salts, phenol, guanidine) | about 1.8 to 2.2 | about 1.8 to 2.2 |
| Integrity | Clear high-molecular-weight band, no smear | RIN 7 or higher for standard poly-A RNA-seq (8+ is safer); DV200 for FFPE RNA |
A low A260/230 is the classic warning sign of leftover salt, phenol or guanidine from the extraction kit. These chemicals also inhibit the enzymes used in library prep, so a sample can pass on amount and still fail later.
Want to check your own sample? Enter your numbers in the free Sample QC checker and see which ones need fixing before you ship.
The one-page Sample QC cheat sheet
Every target on this page on a single sheet you can print and keep next to the Qubit. No signup.
03Send more than the minimum
Send about 1.5 to 2 times the minimum amount the provider asks for. Then they can repeat QC or library prep without asking you for new material, which can save you weeks.
04Use the right buffer and tube
- DNA: 10 mM Tris (pH 8) or the kit's elution buffer. Avoid high-EDTA TE unless the provider allows it, because EDTA can inhibit library prep enzymes.
- RNA: nuclease-free water or the kit's RNA buffer. Don't add carrier RNA unless the provider says it's OK.
- RNA samples: DNase-treat them, or genomic DNA can end up in your RNA library.
- Tubes: use exactly the tube or plate type the provider asks for, and seal plates well.
05Labels that survive dry ice
Use short IDs without spaces or special characters: CTRL_01, not Control #1 (mouse A). Use solvent- and freezer-resistant labels and write on the side of the tube, not only the lid. The ID on every tube must exactly match the sample sheet. Mismatches are a common cause of delays.
06Ship so samples arrive frozen
- Ship Monday to Wednesday, so the box doesn't sit in a warehouse over the weekend.
- Pack enough dry ice for 2 to 3 extra days in case customs holds the package.
- For international shipments, ask the provider for their customs documents before you ship.
- Email the tracking number and the sample sheet to the provider on the day you ship.
Before you ship: a quick checklist
- I checked the provider's sample requirements for my method
- Amount measured by fluorometer, 1.5 to 2 times the minimum
- Purity ratios and integrity are within range
- Correct buffer and tube type
- Labels are durable and match the sample sheet exactly
- Enough dry ice, shipping Monday to Wednesday, customs documents included
- Tracking number and sample sheet emailed to the provider
Common questions
My RNA has a RIN of 6.5. Can I still sequence it?
Maybe, but ask your provider first. Standard poly-A RNA-seq usually wants RIN 7 or higher. rRNA depletion copes better with partly degraded RNA. Also check that RIN is similar across your groups, so degradation doesn't look like biology.
Why did the provider measure less DNA than I did?
Usually because you measured with a NanoDrop and they used a fluorometer. The NanoDrop also counts RNA, free nucleotides and some contaminants.
Do I need to DNase-treat RNA before sequencing?
Yes, unless your provider does it. Leftover genomic DNA ends up as background reads in your RNA library.
Our step-by-step sample QC checklist is also published as a citable protocol on protocols.io: Pre-submission QC checklist for DNA and RNA samples sent for sequencing.
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