Page 38 - IBRO_RNA School_Abstract Book
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Oligo dT Adaptor ligation:
Upon the PolyA tailing process, OligodT adaptor for universal reverse prier
is ligated to the RNA by incubating at 60°C for 10 minutes.
Reverse transcription:
Once the adaptor is ligated, the whole product is reverse transcribed using
reverse transcriptase enzyme and incubating the mix at 42°C for 1hr and
82°C for 10 min
qRT-PCR:
Expression of microRNA is quantifies using SYBR green based quanti-
tative Real-Time Polymerase Chain Reaction (qRT-PCR). cDNA is used
as template and Forward primer specific to our miRNA of interest is used
paired with a Universal Reverse Primer. Cycle threshold value (ct-value) is
used to quantify the expression level.
Ct-value: Number of PCR cycles required to attain a threshold level.
