Total RNA was extracted from three 3 mm punches of NBS using the illustra RNAspin Mini RNA kit (GE Healthcare). Due to large sample size, all spots were not extracted at the same time but in batches of 24. Extracted mRNA was treated the same way for all spots prior to analysis. To minimize variables, all assays were performed by a single technician and extracted mRNA was immediately stored at −80 °C until the next procedure. The Agilent Whole Human Genome Gene Expression 8 × 60 K Microarray platform was used for microarray assays. Quantitative real-time PCR (qRT-PCR) was performed using Applied Biosystems 7500 Fast Real-Time PCR System. Details regarding sample processing, RNA isolation, microarray and qRT-PCR techniques were published in our previous study [8 (link)].
Illustra rnaspin mini rna kit
Manufactured by GE Healthcare
The Illustra RNAspin Mini RNA kit is a lab equipment product designed for the purification of RNA from a variety of sample types. It provides a simple and efficient method for extracting high-quality RNA for use in downstream applications.
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2 protocols using illustra rnaspin mini rna kit
1
Microarray and qRT-PCR Analysis of NBS
2
Total RNA Isolation and Quality Assessment
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Total RNA was isolated from the right tibias using the TRIzol ® reagent (Invitrogen, Carlsbad, California) according to the manufacturer's instructions. After the RNA isolation, the samples were purified using the illustra RNAspin Mini RNA Kit (GE Healthcare Life Sciences) according to the manufacturer's instructions, quantified by NanoVue spectrophotometer (GE Healthcare Life Sciences). The quality and integrity of the total RNA were confirmed with an Agilent 2100 Bioanalyzer (GE Healthcare Life Sciences) and samples presenting RNA integrity numbered ≥8 were used for cRNA synthesis.
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