FastLane Cell Multiplex Kit (200)

PCR Multiplex PCR - Mammalian DNA

Experiment
PCR Multiplex PCR - Mammalian DNA
Product
FastLane Cell Multiplex Kit (200) from Qiagen
Manufacturer
Qiagen

Protocol tips

Protocol tips
Follow manufacturer’s instructions

Publication protocol

Monocyte-derived macrophages from each animal were seeded, in triplicates, in a 96-well plate. Each well was seeded with 100 µL cell suspension containing 40,000 cells in the growth medium. After incubation for 48 h, they were induced with 1 µg/ml lipoprotein Pam3CSK4 (Invivogen cat No. tlrl-pms). Cells in control wells were kept uninduced. Six h after induction, cells were processed for real-time, one-step RT-PCR using a FastLane Cell Multiplex Kit (Qiagen cat No. 216513) following the protocol recommended by the manufacturer. Briefly, media from the wells were aspirated and to each well 125 µL of Buffer FCW was added. Immediately after this, Buffer FCW was removed and to each well 50 µL of cell processing mix was added and the plate was incubated at room temperature for 5 min. The FastLane cell lysates containing stabilized RNA obtained in this way from each well were then transferred to 0.6 ml centrifuge tubes and incubated for 5 min at 75°C. The concentration of the RNA was determined using Qubit RNA Assay Kit (Invitrogen cat No. Q32852) and Qubit 2.0 Fluorometer (Invitrogen cat No. Q32866). Finally, the concentration of all the samples was adjusted to 20 ng/µL.

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Reviews

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Discussion

Discussion

5 years ago

Author: R. Verma India

Multiplex PCR and qPCR

I have been doing multiplex PCR and qPCR separately. Is there a way to combine them in a single reaction using Sybr Green?

Discussion

5 years ago

Author: Roland Lafontaine France

My multiplex PCR has decreased in sensitivity

Hello there! My multiplex PCR has decreased in sensitivity even though I have not changed any of the parameters. The materials are in good condition and the PCR machine is well calibrated. I have tested multiple samples but one of my targets has decreased in sensitivity.

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Papers

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Manufacturer protocol

Download the product protocol from Qiagen for FastLane Cell Multiplex Kit (200) below.

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