GSM8634034: sip53-1; Homo sapiens; RNA-Seq
RNA is extracted from the sample using a reagent like TRIzol. The quality and quantity of RNA are assessed using a spectrophotometer or a bioanalyzer. Next, the RNA is typically treated with DNase to remove any contaminating genomic DNA. Poly(A) RNA is then enriched, often using oligo(dT) beads, to isolate mRNA from the total RNA pool, although ribosomal RNA (rRNA) depletion methods can also be used. The mRNA is then reverse transcribed into cDNA using a reverse transcriptase enzyme. The cDNA is fragmented to a suitable size, and adapters are ligated to the fragments to facilitate amplification and sequencing. The library is amplified by PCR, and the resulting products are sequenced using high-throughput sequencing platforms (e.g., Illumina, PacBio, or Oxford Nanopore). The data are then analyzed to quantify gene expression, detect novel transcripts, and explore alternative splicing events. After amplification, the library is typically size-selected to ensure a uniform fragment size distribution, and the quality and quantity of the library are assessed using techniques such as gel electrophoresis or a Bioanalyzer. The final library is ready for sequencing on a high-throughput platform.