Please remember that it is your responsibility to keep track of the identification of your sample. When filling out the Sample Submission Form, use a sample ID and description that matches your records. Please refrain from using duplicate identifiers from previously submitted samples to avoid confusion.
Samples must be in nuclease-free 1.5-2.0 mL Eppendorf Safe-Lock microcentrifuge tubes.
Evaluate quantity of samples by fluorometry. Spectrophotometric based methods tend to overestimate sample concentration, resulting in insufficient starting material.
DNA/RNA should be high quality with an OD 260/280 of 1.8-2.0 and an OD 260/230 of 2.0-2.2.
Prior to submitting samples, it is required to submit all quality control measurements performed. Our staff will review your data and advise you on the suitability of your samples for satisfactory Illumina sequencing.
If you do not have access to proper quality control instruments, the GCF will quantify and test the integrity of all samples before accepting them for sequencing.
Project work will begin once a signed quote, Sample Submission Form, and all quality control data have been received by the GCF.
The GCF recommends at least n=3 biological replicates be submitted for each sequencing experiment. Samples may exhibit biological variation and replicates are statistically vital for downstream data analyses.
Resuspend DNA samples in nuclease-free water or 10 mM Tris-HCl, pH 7.0-8.5. Avoid DEPC treated water and resuspension buffers containing detergents, as they could interfere with library preparation and/or sequencing.
Provide at least 500 ng of DNA with a concentration of ≥ 50 ng/µL.
Evaluate DNA integrity by gel electrophoresis or automated electrophoresis using instrumentation such as the Agilent Bioanalyzer or TapeStation. If using gel electrophoresis, high quality genomic DNA should give a major band of 10-20 kb. Bioanalyzer or TapeStation data is preferred.
Always wear gloves when working with RNA and keep all samples on ice while processing.
Resuspend RNA samples in RNAase-free water or 10 mM Tris-HCl, pH 7.0-8.5. Avoid DEPC treated water and resuspension buffers containing detergents, as they could interfere with library preparation and/or sequencing.
Provide at least 500 ng of RNA with a concentration of ≥ 50 ng/µL.
Submit high quality RNA with a RIN (RNA Integrity Number) or RINᵉ (RNA Integrity Number Equivalent) value of ≥ 7 as assessed by automated electrophoresis using an Agilent Bioanalyzer or TapeStation. If using gel electrophoresis, high quality RNA will exhibit a 2:1 ratio of 28S:18S rRNA. Bioanalyzer or TapeStation data is preferred.
Provide ≥ 20 µL of your undiluted 10 nM library suspended in 10 mM Tris-HCl, pH 7.0-8.5.
If you have custom primers, please provide ≥ 40 µL (10 nM) of your primers with your libraries.
Quantify libraries using qPCR.
Evaluate purity of libraries using a spectrophotometric method.
Evaluate quality of libraries by gel or automated electrophoresis using instrumentation such as the Agilent Bioanalyzer or TapeStation. Bioanalyzer or TapeStation data is preferred.
The GCF strongly recommends that library preparation be performed by GCF personnel.