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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.

 

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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.

 

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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).

 

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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).

 

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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).

 

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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.

 
 

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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