Department of Botany & Plant Sciences

Best Practices for Illumina Library Submission

Best Practices Illumina Library submissions

  1. For most libraries, the core needs a minimum of 10 ul of library (or library cocktail) at 5 nM concentration. Higher library concentration and volume would be encouraged but it is unnecessary to submit the entire library thus the core will have sufficient volume for the sequencing run and determine the quality of the library. Sequencing libraries with a concentration of lower than 5 nM is possible. Please discuss with core personnel. 
  1. Library/cocktail concentration should be determined by Qubit prior to submission for accurate quantification.
  1. Please insure that all libraries in a cocktail are of similar lengths and verify that the when sequencing the expected output length will not go too far into the adapter region.
  1. If custom sequencing primers are required for library sequencing, please provide custom primers at the time of library submission, along with their concentrations.
  1. Please refrain from using any spaces or special characters in your sample and/or barcode names. Numbers, letters, dashes, and underscores only please.
  1. If submitting a spreadsheet of your barcodes that has both the barcode sequence and the reverse complement sequence, please indicate clearly which is to be used for sequence analysis.
  1. Any existing knowledge you have of issues with your library (low diversity, poor cluster generation, etc.) can be very useful and should be mentioned. Similarities to previous libraries run can also be extremely useful and should be mentioned.
  1. We strongly suggest doing a bead cleanup on the final library before submitting it for sequencing in an effort to remove adapter dimers and contaminants which can negatively affect cluster efficiency and ultimately data generation.
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