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Frequently Asked Questions

Sample Prep & Sequencing: Frequently Asked Questions

Sample Submission

What types of samples are accepted?

We accept DNA and RNA samples that are plated in our designated barcoded plates.

What are the DNA requirements for submission?

Typically, we request 3ug high-quality DNA from each individual in the project. However, we do accept samples with much less total mass on a case by case basis. Samples should be provided as 30uL normalized to 100ng/uL in 1xTE buffer. Upon request, we can return any unused DNA to the investigator at the end of the project.

What if I don’t have 3ug of DNA for each participant and there is no possibility to re-collect samples for my project?

In some circumstances, we may approve a project for low-input exome or genome sequencing.

I have samples that I want to submit for my project, but some are below the required concentration of 100ng/uL? Can I still submit them?

If you have sufficient DNA mass but the samples are too dilute, the samples may be concentrated 2-3 fold in a speedvac to bring the concentrations into the required range. The final volume for each sample must still be at least 30uL, which is the minimum volume required for submission.

Can I suspend the DNA in TE buffer with EDTA at 1mM concentration instead of the 0.1 mM requested?

No, please use the recommended buffer. We prefer to use this concentration to minimize chelation that can interfere with some of the library prep reactions.

When submitting a manifests for multiple plates, can I submit one spreadsheet, but put the different plates on separate tabs?

No, please submit one manifest spreadsheet per DNA plate.

Can I use subjects first and last names as sample IDs?

No, do not use subject names. Please do not send any information that links to subject identity.

Can I alter the submission manifest?

You may add columns, but do not remove columns or change the names of any of the columns.

There are hidden rows in the manifest, may I remove them?

No, those are hidden rows that are needed for entering the manifest into our LIMS.

May I send my samples at room temperature?

Since we do not accept samples in screw-top lids, please do not send samples at room temperature. Samples that are packed frozen and that stay frozen during delivery are much less likely to contaminate each other.

I am in the Seattle area. May I pick up my plate and bring it back when I’ve aliquoted out my DNA?

Yes, please feel free to drop by the lab. We can send instructions for how to reach us.

QC

What happens to samples that fail QC?

Samples that fail QC are not eligible to proceed into the sequencing pipeline. A single sample may be excluded from the project or in cases where the failing sample is critical to the analysis (i.e. the proband in a trio) the entire family will be excluded.

I’ve received notification that one or more of my samples has failed QC?

If a clerical error on the manifest has caused the sample to fail the sex check, you may notify the NWGC of the need to correct a manifest. If a sample has failed due to a sex conflict or due to insufficient concentration, you may submit a replacement sample by filling out a new manifest describing the replacement sample. Submission of replacement samples will delay the release of data for your project.

Sequencing

What is the turnaround time for sequencing?

It typically takes 12 weeks after all samples in the project pass QC for data to be released. Projects with a large number of samples may require extra time for sequencing before all samples are finished.

Data Release

How will I know when my data are available?

You will receive an email notifying you of data release.

What data do I get back from the NWGC?

You will receive the following files: bam, bam index, multi-sample vcf, sample ID lookup table, genotype data (if applicable), and SeattleSeq annotation.

Can the UW-CMG provide FASTQ files in addition to bam files?

You can generate FASTQ files from the BAMs received in data release. Instructions.

Can I get additional bioinformatics help on my project?

Yes, our bioinformatics team is available to assist at an additional hourly rate.

Sequencing Only: Frequently Asked Questions

Can I submit custom primers with my samples?

Yes, if necessary. The amount will vary depending on the platform used, but typically 20uL of 100uM primer is sufficient.

I do not need a lot of data, can I order part of a lane?

No, we do not sell partial lanes. We recommend either choosing another platform that generates less reads or waiting until you have more samples to submit.

When will I receive my data?

Our typical turnaround time is 4 weeks from submission to data being ready for download. This time may be shorter or longer depending on our current workload.

In what format will I receive my data?

Raw run data will be converted to .fastq format, then put on our server for you to download.

Genotyping Only: Frequently Asked Questions

What happens to samples that fail QC?

Samples that fail QC are not eligible to proceed. A single sample may be excluded from the project or in cases where the failing sample is critical to the analysis (i.e. the proband in a trio) the entire family will be excluded.

I’ve received notification that one or more of my samples has failed QC?

If a clerical error on the manifest has caused the sample to fail the sex check, you may notify the NWGC of the need to correct a manifest. If a sample has failed due to a sex conflict or due to insufficient concentration, you may submit a replacement sample by filling out a new manifest describing the replacement sample. Submission of replacement samples will delay the release of data for your project.

In what format will I receive my results?

Our project managers will work with you to generate a custom final report for your samples. Raw data files may also be sent.