Links
- UCSF CAT Services Requests iLabs
- UCSF iLabs account creation guide
- CAT Sequencing Queue
FAQ
Q: How do I submit my sample?
A: Fill out the iLabs service request and submission form, attach(upload) your bioanalyzer traces, and then drop off the sample to our lab. See the complete instructions for Illumina here and pacbio here.
Q: Can I reserve a spot in the queue?
A: No, we do not reserve spots in our queue. Samples are put in the queue after we have received the submission; however, if your sample tube is not received before its assigned flow cell is ready to process, your sample will be dropped to the bottom of the queue.
Q: Do I really have to run my Illumina library on the Bioanalyzer?
A: It's not a requirement but good practice to QC your Illumina sample. From the Bioanalyzer (or similar instruments), we use the average size of the Illumina library peak for QC purposes, and it can also be used to identify any adapter dimers, so for accurate quantitation, yes, but at the very least, know your average size and write it in during the submission process. Without this information, we will not be able to get accurate clustering. You should always QC your Illumina libraries before submission. Illumina libraries with free primers and adapter dimers will not sequence well; this is especially true for the NovaSeq X, where there is a strong bias towards smaller library molecules.
Q: What concentration and volume should my Illumina library be for submission?
A: See the complete instructions for Illumina here and pacbio here. A volume of 20ul of 5-10nM is ideal. If your library is lower than 5nM, please increase the volume to 30ul. If your sample is greater than 20 nM, please dilute it; otherwise, we may have to repeat quantitation, which will delay your results.
Q: What is the cost to sequence at the CAT?
A: Please see the sequencing page for the costs associated with each type of sequencing run configuration that we offer.
Q: What if I have a custom reads configuration and/or custom primer?
A: Since both sequencing read configuration and primers are applied to the whole flow cell, you will need to run a full-flow cell. It is simpler if all Illumina libraries are made with adapters compatible with the Illumina sequencing primers; however, if you still choose to use a custom sequencing primer, then we ask that you clearly indicate that in the special notes section of the submission and provide an aliquot, 10ul of a 100uM aliquot in a 1.5ml tube. You are responsible for failed runs caused by bad custom primers.
Q: When will my sample be run?
A: When a sample library will be run is dependent on the length of the queue: how many samples have already been submitted for the same run configuration and when the run configuration's flow cell fills up. You can check on where your library falls or how close the flow cell is to filling up by looking at the queue page. Samples currently on the sequencing are in bold. Requests or emails inquiring about when your sample will be run are, as a policy of the CAT Core, never answered, we provide the queue as a means of being able to estimate when your sample may be run.
Q: Can I get my samples back after they are run?
A: The CAT will keep your sample viable for 3 months; please contact the CAT staff to request your sample back. We suggest only submitting the needed aliquot of your library and keeping the rest for yourself.
Q: I didn’t get enough data the first time, can I rerun my library?
A: We can rerun a sample if we will still have the sample in storage, and if the remaining amount has enough volume left for a second run. We will need a new submission form filled out for the rerun of the library.
Q: How do I download my data?
A: When your data is ready, you will receive a "Data Ready" e-mail through iLabs with instructions on how to access our sFTP server. You are also able to download data via the UNIX rsync command. Data is available for approximately 3 months on a fast solid-state array storage server, before being moved to a slower disk array for approximately another 3 months. Please download and backup your data as soon as possible.
Q: How do I use sFTP?
A: Below is a video showing how to log in and grab your data, Cyberduck is a free program for Windows and Mac OS that is easy to use. Note that our new server address is fastq.ucsf.edu.
Q: I've lost my data. Do you still have it, and can I get a copy?
A: We have a finite amount of storage space, and our lab generates a significant amount of data. We only guarantee to store data for retrieval for 6 months; at which point, it is only kept until we need room to store more recent runs. We suggest backing it up in multiple places after copying it from us.
Q: What form does the data come in?
A: Illumina data is in compressed FASTQ files, split by the barcode/index provided in the submission. PacBio data is in compressed sam (.bam) files, split by the barcode/index provided in the submission.