Bioanalyzer hs dna assay kit

Single-cell libraries were constructed using the 10X Genomics Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 with Single Index Kit T Set A. Due to the extremely low RNA content of single-cell malaria parasite and the AT-rich genome (~70% AT), modifications to the cDNA amplification and library preparation workflow were made accordingly. These modifications included - 30x cDNA amplification cycles; taking 50% cDNA as input into library generation; reducing fragmentation time to 2 minutes; and changing the extension time to 65 °C during index PCR. Individual library Qc was performed using the BioAnalyzer HS DNA Assay kit (Agilent).

Library concentration was determined with the KAPA Library Quantification Kit (ROCHE) using the QuantStudio3 Real-Time PCR systems (ThermoFisher) and assessed for fragment size using the BioAnalyzer HS DNA Assay kit (Agilent). Following library pooling in equimolar concentrations, a total of 1.2 nM library was sequenced on the Illumina NovaSeq 6000 with SP flow cell using version 1.5 chemistry as follows: Read 1 – 28bp; Index read i7 – 8bp; Index read i5 – 0bp; and Read 2 – 91bp.

Single-cell libraries were constructed using the 10X Genomics Chromium Next GEM Single Cell 3′ Reagent Kits v3.1 with Single Index Kit T Set A. Due to the extremely low RNA content of the single-cell malaria parasite and the AT-rich genome (~70% AT), modifications to the cDNA amplification and library preparation workflow were made accordingly. These modifications included: 30× cDNA amplification cycles, taking 50% cDNA as input into library generation, reducing fragmentation time to 2 min, and changing the extension time to 65 °C during index PCR. Individual library quality control was performed using the BioAnalyzer HS DNA Assay kit (Agilent).