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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    
 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)    
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    
 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
  1. M-MSP methylated MSP, U-MSP unmethylated MSP, Tm annealing temperature, SRBS seed region binding site