AVITI and AVITI24 Enhanced Output Datasets
AVITI™ and AVITI24™ were used to perform sequencing of WGS, RNA-Seq, scRNA-seq and Trinity™ libraries. Additional details are available in the Enhanced Output with AVITI24 tech note.
Methods
Trinity
Libraries were prepared for Trinity sequencing with the Twist for Element Exome 2.0+ Comprehensive Exome Spike-in Library Prep and Standard Hybridization with Trinity and the xGen Exome Sequencing Kit Trinity for Element AVITI System protocols. Each library prep used 100ng of HG001 as input and Elevate full-length adapters. Twist libraries were hybridized in a pool of 2 µg total. IDT xGen libraries were hybridized in a pool of 4 µg total. All other standard hybridization procedures were followed according to the above protocols. 36 samples were sequenced for each protocol on the AVITI and AVITI24 with Trinity and a 2x150 read length. Bases2fastq was used to generate FASTQ files.
Whole-Genome Sequencing
PCR-free libraries were prepared with the Element Elevate™ Mechanical Fragmentation Library Prep protocol and kit with full-length Elevate adapters. Libraries were prepared with 1 µg of HG002, HG003, or HG004 DNA. A 0.46x/0.62x double-sided SPRI was used for size selection. Libraries were sequenced on the AVITI and AVITI24 systems with Cloudbreak Freestyle and a 2x150 read length. Bases2fastq was used to generate FASTQ files.
scRNA-Seq
Single-cell RNA-Seq libraries were prepared with the 10X Chromium Single Cell 3’ Gene Expression library kit using ~10,000 human PBMC cells. Libraries were end-polished before sequencing on AVITI and AVITI24 with Cloudbreak Freestyle and a 28+90 cycle read length. Bases2fastq was used to generate FASTQ files with the legacy-fastq naming convention applied.
RNA-Seq
RNA-Seq libraries were prepared with the KAPA HyperPrep mRNA kit (Roche) using 100ng of either Universal Human Reference RNA (Thermo Fisher) with ERCC RNA spike-in mix 1 (Thermo Fisher) or Human Brain Reference RNA (Thermo Fisher) with ERCC spike-in mix 2. Five replicates of each sample were pooled after library prep for sequencing. Libraries were sequenced on the AVITI and AVITI24 with Cloudbreak Freestyle and a 2x75 read length. Bases2fastq was used to generate FASTQ files.