To prepare for the miRNA microarray analysis, the reaction was prepared using miRNA complete Labeling and Hyb kit (24X; Agilent Technologies, Inc.; cat. no. 5190-0456) through dephosphorylation, denaturation and ligation, according to Agilent miRNA microarray protocol. Subsequently, the labelled RNA was purified using Micro Bio-Spin 6 (Bio-Rad Laboratories, Inc.; cat. no. 732-6221) and dried using a vacuum concentrator at 45–55°C or on the medium-high heat setting. After preparing the hybridization samples, hybridization was conducted in the Agilent G2545A Hybridization Oven (Agilent Technologies, Inc.) at 55°C for 20 h. In addition, the microarray slides were washed, scanned and finally the images were added to be extracted using the Agilent Feature Extraction software v10.5.1.1 (Agilent Technologies, Inc.) to analyze and generate the miRNAs. In order to perform the differential expression analysis, a DEGseq package tool (15 (link)) was applied and the significantly differential miRNAs were identified according to the P-value (<0.05).
Agilent g2545a hybridization oven
Manufactured by Agilent Technologies
Sourced in China
The Agilent G2545A Hybridization Oven is a laboratory equipment used for controlled incubation and hybridization of microarrays or other samples. It provides precise temperature control and mixing capabilities to facilitate various hybridization experiments.
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2 protocols using agilent g2545a hybridization oven
1
miRNA Microarray Analysis Protocol
2
Profiling ceRNA Expression via Microarray
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Total RNA was extracted and purified with PAXgene™ Blood RNA Kit (Cat#762174, QIAGEN, GmBH, Germany) following the manufacturer’s instructions. Qualified RNA was amplified and transcribed into cRNA, which was purified and hybridized to the SBC human ceRNA array V1.0 (Biotechnology Corporation, Shanghai, China) in Agilent G2545A Hybridization Oven. Slides were washed and then scanned by Agilent G2565CA Microarray Scanner with default settings. Raw data were normalized by the Quantile algorithm of the “limma” package of R 3.2.2 software (Ritchie et al., 2015 (link)).
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