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Freshly isolated tissues were prepared into a single-cell suspension, and their quality was examined and counted. The cells were washed, resuspended, and prepared into an appropriate cell concentration for downstream operations on the 10x Genomics Chromium™ system. Based on the expected number of harvested cells, GEMs (Gel Bead in Emulsion) were constructed for single-cell separation. Once the GEMs were formed, they were collected and subjected to reverse transcription for labeling in a PCR machine. Following oil-treatment of the GEMs, the amplified cDNA was purified and enriched using magnetic beads, and subjected to amplification and quality control for library construction. After fragmentation, adapter ligation, and sample index PCR, etc., the library was quantified using Qubit. Sequencing was performed on the Illumina NovaSeq 6000 instrument using the 150-basepair paired-end reads and the generated data was utilized for bioinformatics analysis |
RNA-Seq |
TRANSCRIPTOMIC |
PCR |
SINGLE
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