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This allows us to use the kit for all experiments, reducing variability due to differences between kits.
Use the kit when necessary.
The RNA subsequently went through a clean-up step using the RNeasy Mikro kit from Qiagen.
For epithelial cell lines, these steps were combined using the 2× One-Step RT-ddPCR Kit for Probes (BioRad).
PCR amplification was carried out using the core kit for SYBR Green Applied Biosystemss, Foster City, CA, USA) and the Step One Real Time PCR System (Applied Biosystems).
Subsequent steps were carried out using the Vectastain ABC kit according to manufacturer instructions.
A 1-step RT-PCR was then performed by using the One-Step RT-PCR Kit (QIAGEN).
RT-PCR was performed by using the One-Step PCR Kit (QIAGEN), and the RT-PCR product was used as template for nested PCR.
The cultures were analyzed for apoptotic DNA fragmentation by using the One Step TUNEL Apoptosis Assay Kit (Beyotime, Shanghai, China).
RT-PCR for hemagglutinin 5 (H5) was then performed on the extracted RNA by using the One-Step RT-PCR Kit (QIAGEN) with an H5-specific primer pair.
Please consider using the Foal Diarrhea/Septicemia Kit and Adult Equine Diarrhea Kit for equine submissions.
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