SsoAdvanced™ SYBR^® Green Supermix

SsoAdvanced™ SYBR^® Green supermix provides extreme sensitivity in the detection of a single copy target gene. The cyclin gene was amplified and detected from fivefold serial dilutions of 10 ng–80 pg () and 3.2 pg () human genomic DNA. Cyclin efficiency = 103%, R² = 1 (for 10 ng–80 pg). Inset shows the standard curve for the various dilutions. Cq, quantification cycle; RFU, relative fluorescence units.

Exceptional reproducibility can be achieved with SsoAdvanced™ SYBR^® Green supermix. Efficient discrimination and reliable quantification can be obtained from a 1.33-fold serial dilution of input template. The GAPDH gene was amplified from varying amounts of HeLa cDNA (1 ng–136 pg). From left to right: 1 ng, 753 pg, 565 pg, 425 pg, 320 pg, 240 pg, 181 pg, and 136 pg. GAPDH efficiency = 96.2%, R² = 0.999. Inset is a magnified view showing robust discrimination and reproducible amplification (six replicates for each input amount). RFU, relative fluorescence units.

SsoAdvanced™ SYBR^® Green supermix demonstrates superior inhibitor tolerance. The ADAR gene was amplified using 5 ng human genomic DNA in the presence of 0.5% food inhibitor. The real-time amplification plot includes various SYBR^® Green qPCR mixes. SsoAdvanced™ SYBR^® Green supermix (); reagent I (); the other three competitor reagents showed failed amplification. RFU, relative fluorescence units.

SsoAdvanced™ SYBR^® Green supermix demonstrates superior inhibitor tolerance. The ADAR gene was amplified from HeLa cDNA in the presence of water alone, or in the presence of a known PCR inhibitor, Eagle's minimal essential medium (EMEM) with fetal bovine serum (FBS; 0, 2.5, 5, 10, and 20%), added to SsoAdvanced™ SYBR^® Green supermix () or a traditional Taq DNA polymerase–based qPCR master mix (). SsoAdvanced™ SYBR^® Green supermix showed quality amplification in all reactions (EMEM with 20% FBS data shown), while the Taq DNA polymerase–based qPCR master mix failed to amplify in all EMEM with FBS combinations (shown in the inset melt curve). RFU, relative fluorescence units.

SsoAdvanced™ SYBR^® Green supermix maintains exceptional stability on the CFX automation system. Tenfold serial dilutions of 100 ng–1 pg cDNA from human spleen were used in each 20 µl reaction to detect GAPDH. All reactions were assembled and loaded onto the CFX automation system and run after varying times (0–144 hr) on the CFX96™ real-time PCR detection system. The thermal cycling conditions were 95°C for 30 sec, then 35 cycles of 95°C for 10 sec and 60°C for 30 sec. Results (GAPDH efficiency, R², and slope) for five time points are shown in the table above. The amplification plot shows real-time traces from 0 hr () and 144 hr (). RFU, relative fluorescence units.