title: +H2
design: 2.1 Sample TestingSelect appropriate detection methods for quality control based on sample and product requirements.2.2 mRNA IsolationTake a specified amount of RNA sample, denature at appropriate temperature to unfold secondary structures, and enrich mRNA using oligo(dT) magnetic beads.2.3 mRNA FragmentationAdd fragmentation reagents to the mRNA from the previous step, react at suitable temperature for a defined duration to fragment mRNA.2.4 cDNA SynthesisPrepare first-strand synthesis reaction mixture and set reaction program for first-strand cDNA synthesis; prepare second-strand synthesis reaction mixture and set reaction program for second-strand cDNA synthesis.2.5 End Repair, A-Tailing, and Adapter LigationPrepare reaction mixture and set reaction program to repair double-stranded cDNA ends and add an adenine (A) base to the 3 end; prepare adapter ligation reaction mixture and set reaction program to ligate adapters to cDNA.2.6 PCR AmplificationPrepare PCR reaction mixture and set reaction program to amplify the product.2.7 Library Quality ControlSelect appropriate detection methods for library quality control based on product requirements.2.8 CircularizationDenature PCR products into single-stranded DNA, prepare circularization reaction mixture and set reaction program to generate single-stranded circular products, digest linear DNA that remains uncircularized.2.9 SequencingSingle-stranded circular DNA undergoes rolling circle replication to form DNA nanoballs (DNBs) containing multiple copies. Load DNBs onto high-density DNA nanoarray chips via mesh-like microwells and perform sequencing using combinatorial probe-anchor synthesis (cPAS) technology