RNAseq of human colorectal cancer cell line Caco-2 with expression of NHE9 against empty vector controls
RNAseq of human colorectal cancer cell line Caco-2 with expression of NHE9 against empty vector controls
Cells were collected in the TRIzol reagent and lysed by repetitive pipetting. Cells were incubated at 37Â°C, 5% CO2, and 95% humidity. Empty vector control in Caco-2 cells by lentiviral vector-based transduction.  Qiagen RNeasyMini Kit according to the manufacturer instructions. Total RNA was isolated and sent for RNAseq analysis The RNA quality assessment is done using RNA screen Tape system. RNA concentration is determined in a Qubit.3 Fluorometer using the Qubit RNA BR assay kit. The fragmented mRNAs were primed with random hexamers, and then reverse transcribed to form cDNA. The cDNA fragments obtained were cleaned up by using AM pure beads. The DNA underwent end repair wherein the mix converts the overhangs resulting from fragmentation into blunt ends. The samples were further purified using AMPure beads and The DNA was then enriched by PCR using illumina universal primer and sample-specific octamer primers. The amplified products were cleaned up by using AM pure beads and the final mRNA library was eluted.