Factors that may influence optimal loading concentration include, but are not limited to:

• Average insert size.

• Number of functional molecules within the library, which means inserts with an adapter at each end.

• Quality of the library, for example, no contaminating short fragments.

• Accurate quantitation of the library; qPCR is ideal.

Note

PCR favors the amplification of shorter fragments over longer ones in a complex mixture of different length templates. This differential amplification needs to be taken into consideration to maximize sequencing yield. For example, if your library size range contains a large number of shorter fragments, which can yield extremely bright clusters, consider a lower loading concentration or perform additional size selection, where possible.

To help you determine your loading schedule, see the Loading Concentration Calculator at https://techwriting.singulargenomics.com/calculator/Loading-Conc-Calc/Content/Topics/Calculators/Loading-Concentration-Calculator.htm