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\nQuantification with Hydrolysis Probes
\n\n\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n
ProcedureTime [min]
Prepare PCR mixes10
Pipette into plate15
PCR run50
Total assay time1 h 15 min
\n\n

\nGenotyping with HybProbe Probes
\n\n\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n
ProcedureTime [min]
Prepare PCR mix10
Pipette into plate5
PCR run50
Total assay time1 h 5 min
\n", "Country": "XG", "Code": "Assay Time", "Name": "Assay Time" }, { "Language": "en", "Value": "Real-time PCR control reactions for quantification and melting-curve based genotyping to prove the performance of the LightCycler® 480 System", "Country": "XG", "Code": "Product Characteristics", "Name": "Product Characteristics" }, { "Language": "en", "Value": "
How this Product Works
Experiment A, Quantification
A 136 bp fragment of the human Cyp2C9 gene is amplified from plasmid DNA and detected with a short hydrolysis probe that is labeled with FAM (probe #80 from the Universal ProbeLibrary*). To test the precision of the LightCycler® 480 System, replicates with only 1,000 or 2,000 copies of target DNA per well are distributed throughout the plate and quantified with reference to a row of standards.
As an internal control (to prove absence of PCR inhibition), a small amount (approx. 100 copies) of an artificial DNA template is added to each well. This control is co-amplified with the target DNA. Its amplification is detected simultaneously with a LightCycler® Red 610-labeled hydrolysis probe. The results are displayed in a separate optical channel. The distances between the wavelengths of the two detection channels (483 – 533 and 558 – 610, LightCycler® 480 Instrument I; 465 – 510 and 533 – 610, LightCycler® 480 Instrument II) are high enough that there is no need to use color compensation to correct for crosstalk.
Alternatively, the target amplification can be detected using SYBR Green I. By subsequent melting curve analysis of the PCR product, the specificity of the reaction can be proven.

Experiment B, Genotyping
The same 136 bp fragment of the CyP2C9 gene is amplified from different samples of plasmid DNA. This gene is known to contain a single nucleotide polymorphism (SNP), and various samples included in the experiment contain the wild type sequence, the homozygous point mutation, and heterozygote DNA with wild type and mutant strands. With HybProbe probes for detection, a subsequent melting curve analysis can be used for identification of the different genotypes, because the probe melts off the perfectly matched sequence and the mismatched sequence at different melting temperatures.", "Country": "XG", "Code": "Principle", "Name": "Principle" }, { "Language": "en", "Value": "\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\t\n\t\t\n\t\n
Vial / BottleCapLabelFunction / DescriptionContent
1yellowLightCycler® 480 Control Kit Standard 1, 102 copies/5 μlTarget: wild type plasmid DNA1 vial, >45 μl
2yellowLightCycler® 480 Control Kit Standard 2, 103 copies/5 μl Target: wild type plasmid DNA1 vial, >450 μl
3yellowLightCycler® 480 Control Kit Standard 3, 2 × 103 copies/5 μl Target: wild type plasmid DNA1 vial, >405 μl
4yellowLightCycler® 480 Control Kit Standard 4, 104 copies/5 μlTarget: wild type plasmid DNA1 vial, >45 μl
5yellowLightCycler® 480 Control Kit Standard 5, 105 copies/5 μlTarget: wild type plasmid DNA1 vial, >90 μl
6yellowLightCycler® 480 Control Kit Standard 6, 106 copies/5 μl Target: wild type plasmid DNA1 vial, >45 μl
7yellowLightCycler® 480 Control Kit Standard 7, HeterozygoteTarget: heterozygous plasmid DNA1 vial, >45 μl
8yellowLightCycler® 480 Control Kit Standard 8, MutationTarget: mutant plasmid DNA1 vial, >45 μl
9blueLightCycler® 480 Control Kit Primer Mix, 20x conc.Mix of two target-specific primers1 vial, 255 μl
10redLightCycler® 480 Control Kit Genotyping Probes, 10x conc.\n\t\t\t
    \n\t\t\t\t
  • HybProbe probe mix
  • \n\t\t\t\t
  • Probe 1: Fluorescein-labeled at the 3' end
  • \n\t\t\t\t
  • Probe 2: LightCycler® Red 640-labeled at the 5' end
  • \n\t\t\t
\n\t\t\t
1 vial, 80 μl
11greenLightCycler® 480 Control Kit Quantification Probe, 10x conc.FAM-labeled hydrolysis probe1 vial, 450 μl
12purpleLightCycler® 480 Control Kit Internal Control, 10x conc.Primer, probe, and template mix, containing LightCycler® Red 610-labeled hydrolysis probe for detection of control DNA sequence1 vial, 450 μl
13colorlessLightCycler® 480 Control Kit Water, PCR GradeTo adjust the final reaction volume1 vial, 1,000 μl
14colorlessLightCycler® 480 Control Kit LC 480 Genotyping Master, 5x conc.Ready-to-use hot start PCR Mix. Contains a modified Taq DNA Polymerase, reaction buffer, dNTP mix (with dUTP instead of dTTP), and 15 mM MgCl2.1 vial, 384 μl
15colorlessLightCycler® 480 Control Kit LC 480 Multiplex DNA Master, 5x conc.Contains qPCR Reaction Buffer, AptaTaq Polymerase, dATP, dCTP, dGTP, dUTP, MgCl2, and proprietary additives.2 vials, 880 μl
16colorlessLightCycler® 480 Control Kit LC 480 SYBR® Green I Master, 2x conc.Ready-to-use hot start PCR reaction mix. Contains FastStart Taq DNA Polymerase, reaction buffer, dNTP mix (with dUTP instead of dTTP), SYBR Green I dye, and MgCl2.3 vials, 1,000 μl
\n", "Country": "XG", "Code": "Content", "Name": "Content" }, { "Language": "en", "Value": "The LightCycler® 480 Control Kit is designed to prove the performance of all components of the LightCycler® 480 System, including instrument, software, consumables, generic reagents, and optional devices such as a pipetting robot. The kit is primarily for use with the LC 480 Multiplex DNA Master (vial 15), or alternatively with the LC 480 SYBR® Green I Master (vial 16) for Procedures A, but it can also be used with the LC 480 Genotyping Master (vial 14) for Procedure B.
The test includes two control experiments. Experiment A is for absolute quantification of prediluted standard DNA. Experiment B is used for genotyping samples with a wild type DNA sequence as well as samples with a homozygous or heterozygous point mutation.
The performance of the kit shown in this Instructions for Use is guaranteed only when it is used with the LightCycler® 480 System.
", "Country": "XG", "Code": "Applications", "Name": "Applications" }, { "Language": "en", "Value": "The LightCycler® 480 Control Kit is designed as a tool to confirm real-time PCR and melting curve analysis performance of the LightCycler® 480 Instrument. The kit can be used to verify the LightCycler® 480 System’s speed, accuracy, and intra-well assay reproducibility. SNP analysis with HybProbe probes and quantification with hydrolysis probes in a dual-color setup can both be demonstrated.", "Country": "XG", "Code": "Product Description", "Name": "Product Description" } ] } } ] }

LightCycler® 480 Control Kit

Real-time PCR control reactions for quantification and melting-curve based genotyping to prove the performance of the LightCycler® 480 System

LightCycler<sup><sup>®</sup></sup> 480 Control Kit

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