Lightcycler 480 software release 1

Manufactured by Roche

Sourced in Germany

The LightCycler®480 software release 1.5.0 is a software package for real-time PCR instrumentation. It provides functionality for data acquisition, analysis, and reporting.

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Lab products found in correlation

Lightcycler 480 instrument 2, by Roche (2 mentions) Taqman microrna assay, by Thermo Fisher Scientific (1 mentions) Taqman primer, by Thermo Fisher Scientific (1 mentions) Lightcycler 480 real time pcr system, by Roche (1 mentions) Lightcycler 480 probes master, by Roche (1 mentions) Faststart essential dna green master, by Roche (1 mentions) Taqman rna to cttm 1 step kit, by Thermo Fisher Scientific (1 mentions) Peqgold hp total rna kit, by Avantor (1 mentions) Taqman gene expression assay, by Thermo Fisher Scientific (1 mentions) Rnalater, by Qiagen (1 mentions)

Close spelling variants of this product

LightCycler 480 Software release 1.5.0 SP3 LightCycler 480 I LightCycler 480 LightCycler software LightCycler 1.5 LightCycler

3 protocols using lightcycler 480 software release 1

Validating miRNA Expression by RT-qPCR

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To validate the differential miRNA expression, RT-qPCR was performed as described previously [17 (link)]. First, 10 ng of total RNA was reverse-transcribed using a multiplex approach with TaqMan primers (reference gene: RNU48 (Assay ID: 001006) or RNU44 (Assay ID: 001094), ssc-miR-335-5p (Assay ID: 244560_mat) and hsa-miR-335* (hsa-miR-335-3p, Assay ID:002185); Applied Biosystems, Carlsbad, CA, USA) [70 (link)]. Subsequently, qPCR was done using TaqMan microRNA assays (Applied Biosystems) and a LightCycler®480 Probes Master (Roche Diagnostics GmbH, Mannheim, Germany).
The reactions were run in triplicate on the LightCycler®480 Real-Time PCR system (Roche Diagnostics GmbH). The LightCycler®480 software release 1.5.0 (Roche Diagnostics GmbH) was used to analyze the data. The relative miRNA expression levels were calculated using the formula 2^−ΔCp.

Ong J., van den Berg A., Faiz A., Boudewijn I.M., Timens W., Vermeulen C.J., Oliver B.G., Kok K., Terpstra M.M., van den Berge M., Brandsma C.A, & Kluiver J. (2019). Current Smoking is Associated with Decreased Expression of miR-335-5p in Parenchymal Lung Fibroblasts. International Journal of Molecular Sciences, 20(20), 5176.

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Quantifying Toxoplasma Parasite Loads

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Semiquantitative real-time PCR analyses were performed to determine parasite loads in brains as described previously [66 (link)]. FastStart Essential DNA Green Master (Roche, Grenzach-Wyhlen, Germany) was used with 90 ng genomic DNA in a reaction volume of 20 μL. Triplicate reactions were developed in a LightCycler 480 Instrument II (Roche, Grenzach-Wyhlen, Germany). After an initial activation step (95 °C for 10 min), 45 amplification cycles were run, comprising of denaturation at 95 °C for 15 s, annealing at 60 °C for 15 s, and elongation at 72 °C for 15 s. The primers manufactured by Tib MolBiol (Berlin, Germany) were used at a final concentration of 0.3 μM. The specific primer pairs are listed in Table 1 (T. gondii B1 gene, murine argininosuccinate lyase/Asl as a reference). For normalization against Asl, target–reference ratios were calculated with the LightCycler 480 Software release 1.5.0 (Roche, Grenzach-Wyhlen, Germany). The resulting data were further normalized against a control group, i.e., the ileum of WT animals.

Fu T., Znalesniak E.B., Kalinski T., Möhle L., Biswas A., Salm F., Dunay I.R, & Hoffmann W. (2015). TFF Peptides Play a Role in the Immune Response Following Oral Infection of Mice with Toxoplasma Gondii. European Journal of Microbiology & Immunology, 5(3), 221-231.

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RNA Isolation and Gene Expression Analysis

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After removal, tissue samples from brains were immediately transferred to RNA later (QIAgen, Germany). Total RNA was isolated as previously described with the peqGOLD HP Total RNA Kit (peqlab, Germany) including an on-membrane DNase I digestion (peqlab, Germany) [18 (link)].
Relative gene expression was determined similar to previous descriptions [18 (link), 68 (link)] using TaqMan® RNA-to-C_T^TM 1-Step Kit (life technologies, Germany). Reactions were developed in a LightCycler® 480 Instrument II (Roche, Germany). Reverse transcription was performed for 15 min at 48 °C followed by 10 min at 95 °C. Subsequently, 45 amplification cycles were run, comprising of denaturation at 95 °C for 15 s and annealing/elongation at 60 °C for 1 min. TaqMan® Gene Expression Assays (life technologies, Germany) were used for mRNA amplification of HPRT (Mm01545399_m1), IDE (Mm00473077_m1), (IL10, Mm00439616_m1), MMP9 (Mm00442991_m1), NEP (MME, Mm00485028_m1), PSMB8/LMP7 (Mm00440207_m1), PSMB9/LMP2 (Mm00479004_m1) and PSMB10/MECL-1 (Mm00479052_g1). HPRT mRNA expression was chosen as reference for normalization and target/reference ratios were calculated with the LightCycler® 480 Software release 1.5.0 (Roche, Germany). Resulting data were further normalized to values of appropriate control groups.

Möhle L., Israel N., Paarmann K., Krohn M., Pietkiewicz S., Müller A., Lavrik I.N., Buguliskis J.S., Schott B.H., Schlüter D., Gundelfinger E.D., Montag D., Seifert U., Pahnke J, & Dunay I.R. (2016). Chronic Toxoplasma gondii infection enhances β-amyloid phagocytosis and clearance by recruited monocytes. Acta Neuropathologica Communications, 4, 25.

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