LightCycler® Nano System Performance Data
The LightCycler® Nano Instrument looks great on your bench - and produces beautiful data. Like all Roche real-time PCR systems, the LightCycler® Nano System uses excellent thermal control for high reproducibility of results obtained with the 32 samples tested in the same run. High resolution for copy number determination and melting curve analysis is achieved with great speed.
Here is a list of examples to illustrate these performance features. Click the ones that are most relevant for your application, and enjoy the data.
| Topic | Template | Detection Format | Reagent |
| Dynamic range of quantification (2-fold dilutions) | plasmid DNA | Universal ProbeLibrary probe | FastStart Essential DNA Probes Master |
| Dynamic range of quantification (10-fold dilutions) | plasmid DNA | Universal ProbeLibrary probe | FastStart Essential DNA Probes Master |
| Reproducibility of Cq values | human genomic DNA | hydrolysis probe | FastStart Essential DNA Probes Master |
| Endpoint Genotyping | DNA | hydrolysis probe | FastStart Essential DNA Probes Master |
| High Resolution Melting | DNA | ResoLight dye | LightCycler® 480 High Resolution Melting Master |
| Absolute Quantification | DNA | SYBR Green I | FastStart Essential DNA Green Master |
| Homogeneity of Tm values | human genomic DNA | SYBR Green I | FastStart Essential DNA Green Master |
Dynamic range of quantification (2-fold dilutions)
| Purpose of the experiment | Dynamic resolution of gene quantification using Universal ProbeLibrary probes. |
| Procedure | Real-Time PCR using 2-fold dilutions of plasmid DNA (starting from one million copies) as template, and a Universal ProbeLibrary probe for product detection. |
| Observation | The LightCycler®Nano Instrument reliably discriminates template dilutions, even differences by a factor of 2, producing reproducible replicate Cq values. |
| Result | Serial amplification plots of 2-fold dilutions show the high resolution detection and differentiation. |
Dynamic range of quantification (10-fold dilutions)
| Purpose of the experiment | Dynamic resolution of gene quantification using Universal ProbeLibrary probes. |
| Procedure | Real-Time PCR using tenfold dilutions of plasmid DNA (1E0 to 1E9 copies) as template, and a UPL probe for product detection. |
| Observation | All dilutions are positive over the range of dilutions down to single copy detection. The LightCycler® Nano Instrument reliably detects over 10 tenfold dilutions of template. Highly reproducible Cq values are produced . UPL probe results are, as expected, even more precise than those obtained in SYBR Green I assays. |
| Result | Serial amplification plots of Cq values from 10-fold dilutions indicate high PCR efficiency. |
Cq homogeneity using hydrolysis probe
| Purpose of the experiment | Homogeneity of results, independent of block position. |
| Procedure | Real-Time PCR using 1E4 template copies in each position. Beta-globin amplification from human genomic DNA using hydrolysis probe detection. |
| Observation | Excellent intra-run Cq homogeneity. |
| Result | Very low variation of Cq values (SD=0.02). |
Fast, accurate endpoint genotyping
| Purpose of the experiment | Show LightCycler® Nano System capability for SNP endpoint genotyping. |
| Procedure | Genotyping analysis on MDR1 gene fragment, present in 3x 10 replicates. FastStart Essential DNA Probes Master kit was used for amplification and detection with hydrolysis probes. |
| Observation | The LightCycler® Nano System differentiated wild type, heterozygote and mutant genotypes in an unbiased way. |
| Result | Scatter plot discriminates genotypes cleanly (blue=wildtype, green=mutant, red=heterozygote). |
Fast, accurate detection of Class I SNPs using HRM
| Purpose of the experiment | Show LightCycler® Nano System doing high resolution melting analysis of Class I SNPs |
| Procedure | HRM analysis on a 149 bp fragment of the Adr-1 gene with an A to G variation. LightCycler® 480 High Resolution Melting Master, including ResoLight Dye, was used for amplification and detection. |
| Observation | Excellent temperature accuracy of the LightCycler® Nano System differentiates wild type, heterozygote and mutant genotypes in an unbiased way. |
| Result | Normalized melting curves (top), and difference plot (below), discriminate genotypes (blue=wildtype, green=mutant, red=heterozygote). |
Accurate absolute quantification
| Purpose of the experiment | Show LightCycler® Nano System quantifying two different targets in one run using internal standard curves. |
| Procedure | Absolute quantification of two targets (CYP2C9 and CYP219) using internal standard curves. FastStart Essential DNA Green Master used for amplification and detection. |
| Observation | Both targets quantified with high accuracy in a single run. |
| Result | Amplification (A) and standard (B) curves for CYP2C9 (blue) and CYP2C19 (green), and unknowns (red) show accurate, reproducible quantification. |
Tm homogeneity using SYBR Green I Detection
| Purpose of the experiment | Verify homogeneity of Tm results, independent of block position. |
| Procedure | Real-Time PCR using 1E4 template copies in each position. Beta-globin amplification from human genomic DNA using SYBR Green I detection. |
| Observation | Excellent intra-run homogeneity. |
| Result | Very low variation of Tm values |
