Prepared Illumina Library Guidelines

The following guidelines apply to those submitting prepared Illumina libraries.

All General Guidelines must be followed in addition to the guidelines specific to Prepared Illumina libraries.

 

This is an accordion element with a series of buttons that open and close related content panels.

Sample Delivery Vessel - General Guidelines

  • Place in a 96-well skirted, translucent plate (e.g., Eppendorf twin.tec #00030129300).
    • Sample placement should be going down columns (A1,B1,C1…A2,B2,C2…) in contiguous wells (no skipping wells, rows, or columns unless “Blank” samples).
    • Plate should be sealed tightly with one of the following:
      • If shipping on dry ice, seal wells with tight-fitting 8-cap strips (e.g., Thermo Scientific #AB-0265)
      • If delivering from on campus, firmly seal wells with a clear adhesive seal (e.g., Bio-Rad Microseal ‘B’ Film #MSB1001).
    • Samples that are submitted in other containers will be transferred to a 96 well plate at the submitting lab’s expense. Please inquire for cost of transfer.

Sample Naming Conventions - General Guidelines

  • Each sample must be identified with a simple, unique identifier. If names have been used in previous submissions, don’t repeat in subsequent submissions.
  • The following conventions are strictly enforced:
    • 3-20 characters
    • A-Z, a-z, 0-9, dash (-) are the ONLY acceptable characters
    • Space, underscore, punctuation, and special characters are NOT ALLOWED
    • No replication

Sample Labeling Conventions - General Guidelines

  • All tubes and plates must include the following information.
    • PI name
    • Date
    • Sample name (tubes only)

Sample Quantification - General Guidelines

  • Upon sample receipt, the UWBC will quantify the DNA with a fluorometric assay. QC measures done by the UWBC are considered the gold standard, regardless of the measurements done by the submitting lab. Prior to proceeding with samples that fail to meet these requirements, the lab will be informed.

Prepared Illumina Libraries - Specific Guidelines

  1. Follow ALL General Guidelines above.
  2. Sample volume of 10-50 µl with a concentration no greater than 50 ng/µl.
  3. Expect A260/A280 of 1.7-1.9. Ratios that deviate significantly from this suggest that DNA quantification isn’t accurate and your library may not have been successfully prepared.
  4. Only provide index sequences. If index sequences are provided in the incorrect orientation or the wrong indexes are provided, a $115 fee will be charged for time spent reanalyzing.
  5. All libraries that are to be run on the Illumina NovaSeq must have unique dual indexes (UDIs) that are at least 3 bp distance from all UDIs reserved by UWBC DNA Sequencing Facility. Use of UDIs that are commercially available through vendors is recommended.
  6. If requesting 2×150 bp Shared NovaSeq sequencing, indexes must be provided as 10 bp. More information can be found here.