Lightcycler 384 system

Manufactured by Roche

The LightCycler 384 system is a real-time PCR (polymerase chain reaction) instrument designed for high-throughput nucleic acid analysis. It is capable of performing quantitative and qualitative PCR in a 384-well plate format. The system utilizes fluorescence-based detection to monitor the amplification of DNA or RNA targets in real-time during the PCR process.

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Lab products found in correlation

Nanodrop, by Thermo Fisher Scientific (2 mentions) Gdna eliminator column, by Qiagen (1 mentions) Iscript cdna synthesis kit, by Bio-Rad (1 mentions) Qiazol reagent, by Qiagen (1 mentions) Random hexamer, by Thermo Fisher Scientific (1 mentions) Trizol reagent method, by Thermo Fisher Scientific (1 mentions) Rneasy lipid tissue mini kit, by Qiagen (1 mentions)

2 protocols using lightcycler 384 system

Quantitative RT-PCR Analysis of Liver Transcripts

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Total RNA was extracted from liver tissue using Qiazol reagent and RNA‐spin columns (Qiagen, Hilden, Germany). Contaminating genomic DNA was eliminated using gDNA Eliminator Columns (Qiagen). A total of 750 ng RNA was converted to complementary DNA (cDNA) (iScript cDNA synthesis kit; Bio‐Rad, Hercules, CA). Quantitative polymerase chain reactions (PCRs) were conducted in a volume of 10 µL containing cDNA equivalent to 3.75 ng total RNA as template. SYBR Green chemistry (qPCR SYBR Hi‐Rox Green Fluorescein Mix; Bioline, London, United Kingdom) and a LightCycler 384 system (Roche, Basel, Switzerland) were used for real‐time PCR analysis. Data were analyzed with LinReg software and expression was normalized to 36B4.⁽²⁹
⁾ Values are presented relative to the mean expression of the examined transcript in baseline liver specimens. Mean Cq values of the studied genes are reported in Supporting Table S1.

Koelfat K.V., van Mierlo K.M., Lodewick T.M., Bloemen J.G., van der Kroft G., Amygdalos I., Neumann U.P., Dejong C.H., Jansen P.L., Olde Damink S.W, & Schaap F.G. (2021). Bile Salt and FGF19 Signaling in the Early Phase of Human Liver Regeneration. Hepatology Communications, 5(8), 1400-1411.

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Tissue-Specific RNA Extraction and Analysis

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RNA from liver, intestine and muscle was extracted using the TRIzol reagent method (Invitrogen) according to the manufacturer's protocol. RNA from the adipose tissue was extracted using the RNeasy lipid tissue mini kit according to the manufacturer's instructions (Qiagen). The quality and quantity of the RNA were checked on a 1 % agarose gel and a nanodrop (Thermo scientific), respectively. Two μg of RNA was converted to cDNA using the enzyme superscript III reverse transcriptase and random hexamers (Invitrogen). After reverse transcription, the cDNA was diluted 50-fold before being used in RT-qPCR.
RT-qPCR was performed on a LightCycler® 384 system (Roche Diagnostics) according to the protocol described previously (including primer details) (4) . The data normalisation was performed using geNorm (39) . The reference genes eef1a and rna18s were used to calculate the normalisation factor in liver and intestine, while a combination of actb and gapdh was used in muscle and actb and eef1a in adipose tissue.

Lokesh J., Delaygues M., Defaix R., Le Bechec M., Pigot T., Dupont-Nivet M., Kerneis T., Labbé L., Goardon L., Terrier F., Panserat S, & Ricaud K. (2023). Interaction between genetics and inulin affects host metabolism in rainbow trout fed a sustainable all plant-based diet. The British journal of nutrition, 130(7).

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