Table 1 Primer sequences and PCR reaction conditions
From: Epigenetic silencing of miR-340-5p in multiple myeloma: mechanisms and prognostic impact
Primer set | Forward primer (5′–3′) | Reverse primer (5′–3′) | Product size (bp) | MgCl2/Tm/cycles | Reference |
|---|---|---|---|---|---|
(I) Methylation-specific PCR (MSP) of RNF130/miR-340 | Â | Â | Â | ||
 M-MSP | TTT CGT AAA TTT TTC GGG TTT TAC | CCG CTA ATC TAA CGA CAA CG | 184 | 2.0 mM/58 °C/38 | – |
 U-MSP | TTT TGT AAA TTT TTT GGG TTT TAT | CCC CAC TAA TCT AAC AAC AAC A | 186 | 2.0 mM/56 °C/38 | – |
(II) Quantitative reverse transcription PCR (qRT-PCR) | Â | Â | Â | ||
 MDM2 | GTC GGA AAG ATG GAG CAA G | TGT GAG GTG GTT ACA GCA | 311 | 60 °C/40 | – |
 NFKB1 | GGC AGC ACT ACT TCT TGA CC | CAG CAA ACA TGG CAG GCT AT | 88 | 60 °C/40 | [58] |
 NRAS | CGC ACT GAC AAT CCA GCT AA | TCG CCT GTC CTC ATG TAT TG | 177 | 60 °C/40 | [30] |
 GAPDH | ACC ACA GTC CAT GCC ATC ACT | TCC ACC ACC CTG TTG CTG TA | 452 | 60 °C/40 | – |
(III) Cloning of luciferase reporter constructs with wild-type SRBSs of miR-340-5p | Â | Â | Â | ||
 SRBS 1 | AAG GGC TAG CTG TGG TTT CTC TTC GGG GA | AGG GTC GAC GTT TTG CAG GCG TTT GAA CA | 214 | 1.5 mM/60 °C/40 | – |
 SRBS 2 | AAG GGC TAG CTG GGA CAT AGT TTG AAG GTG A | AGG GTC GAC TGG ATG CAA AAC CCA CAG AT | 213 | 2.0 mM/56 °C/35 | – |