For each experiment, two biological replicates were used. Samples from different conditions were processed together to prevent batch effects. Total RNA was extracted using the Zymo Direct-Zol RNA MiniPrep Kit (Zymo research) and the miRNeasy Kit with TRIzol protocol and in-column deoxyribonuclease treatment (QIAGEN), respectively. Quality of total RNA was assessed by the RNA integrity number (RIN) using Agilent Bioanalyzer. All retained RNA samples had a RIN > 8. Ribodepletion was performed with the QIaseq FastSelect RNA Removal Kit. Stranded libraries were produced with the KAPA RNA HyperPrep Kit and sequenced on Illumina NextSeq 500.
In column deoxyribonuclease treatment
Manufactured by Qiagen
In-column deoxyribonuclease treatment is a laboratory technique used to remove DNA contamination from RNA samples. It involves the use of a specific enzyme that digests and eliminates DNA molecules within the sample, while leaving the desired RNA intact.
Automatically generated - may contain errors
Lab products found in correlation
Kapa rna hyperprep kit, by Illumina (1 mentions)
Zymo direct zol rna miniprep kit, by Zymo Research (1 mentions)
Nextseq 500, by Illumina (1 mentions)
Agilent bioanalyzer, by Agilent Technologies (1 mentions)
Mirneasy kit, by Qiagen (1 mentions)
Nextseq 500 system, by Illumina (1 mentions)
Rneasy mini kit, by Qiagen (1 mentions)
2 protocols using in column deoxyribonuclease treatment
1
High-quality RNA Extraction and Sequencing
2
Transcriptomic Analysis of Mouse Brain Regions
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The cortex, hippocampus, and cerebellum were dissected from wild-type and CD mice. RNA was extracted using the RNeasy mini kit with in-column deoxyribonuclease treatment (QIAGEN) according to the manufacturer’s instructions. Thirty nanograms of RNA was used as input for preparing 3′ RNA-seq libraries following CelSeq2 protocol (83 (link)), changing the unique molecular identifier to six bases. Sequencing was performed on Illumina NextSeq 500 system.
Raw reads were aligned to the Mus musculus genome (version mm10) using STAR (84 (link)). Reads were quantified using End Sequence Analysis Toolkit (85 (link)) for 3′ RNA libraries. Differential gene expression analysis was done with edgeR (86 (link)). We excluded from the analysis genes that do not reach 5 cpm in at least two samples. Reads were normalized by the trimmed mean method for each experimental group before differential expression analysis. The RNA-seq data have been deposited in Gene Expression Omnibus (GEO) with the dataset identifier GSE246304.
Raw reads were aligned to the Mus musculus genome (version mm10) using STAR (84 (link)). Reads were quantified using End Sequence Analysis Toolkit (85 (link)) for 3′ RNA libraries. Differential gene expression analysis was done with edgeR (86 (link)). We excluded from the analysis genes that do not reach 5 cpm in at least two samples. Reads were normalized by the trimmed mean method for each experimental group before differential expression analysis. The RNA-seq data have been deposited in Gene Expression Omnibus (GEO) with the dataset identifier GSE246304.
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