EpiNext 5-mC RNA Bisulfite-Seq Easy Kit
(Illumina)

EpiNext 5mC RNA 甲基化定序建庫套組

5-methylcytosine (5-mC) in DNA occurs by the covalent addition of a methyl group at the 5-carbon of the cytosine ring by DNA methyltransferases. This process has been well studied and is generally associated with repression of gene expression. It was also observed that in humans, 5-mC occurs in various RNA molecules including tRNAs, rRNAs, mRNAs and non-coding RNAs (ncRNAs). At least 10,275 5-mC candidate sites were discovered in mRNAs and ncRNAs, which cover 10.6% of the total cytosine residues in the transcriptome. 5-mC seems to be enriched in some classes of ncRNA, but relatively depleted in mRNAs. However, the majority (83%) of their candidate sites were found in mRNAs. Within these transcripts 5-mC appears to be depleted within protein coding sequences, but enriched in 5’ and 3’ UTRs. Two different methyltransferases, NSUN2 and Dnmt2 are known to catalyze 5-mC modification in eukaryotic RNA. Recent data strongly suggest that RNA cytosine methylation affects the regulation of various biological processes such as RNA stability and mRNA translation. Furthermore, loss of 5-mC in vault RNAs causes aberrant processing into Argonaute-associated small RNA fragments that can function as microRNAs. Thus, impaired processing of vault ncRNA may contribute to the etiology of human disorders related to NSun2-defciency.

The EpiNext™ 5-mC RNA Bisulfite-Seq Easy Kit (Illumina) is a complete set of optimized reagents designed for easily carrying out RNA bisulfite conversion, followed by a "post-bisulfite" library preparation process for Illumina platform-based bisulfite sequencing, all in one kit. Intended applications include whole transcriptome RNA bisulfite sequencing and various other RNA bisulfite-based next generation sequencing techniques for 5-methylcytosine (5-mC) based RNA methylation analysis. The optimized protocol and components of the kit allow the RNA to be bisulfite converted and fragmented simultaneously, followed by quick non-barcoded (singleplexed) and barcoded (multiplexed) library construction using low-nanogram quantities of bisulfite converted RNA.

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實驗原理

This kit includes all reagents required for a successful RNA bisulfite conversion and bisulfite RNA library preparation using bisulfite-converted RNA generated from a wide range of input RNA amounts (5 ng to 500 ng). In this preparation, RNA is simultaneously bisulfite converted and fragmented to the appropriate length during the bisulfite process. The bisulfite-treated RNA, which is in single stranded form, is then simultaneously converted to double stranded cDNA and adaptor tagged. The tagged fragments are size selected and purified using MQ binding beads, followed by amplification with a high-fidelity PCR Mix, which ensures maximum yields from minimum amounts of starting material and provides highly accurate amplification of library cDNA with low error rates and minimum bias.

Workflow of the EpiNext™ 5-mC RNA Bisulfite-Seq Easy Kit (Illumina).

Size distribution of library fragments: Post-bisulfite cDNA library was prepared from 50 ng of input mouse RNA using the EpiNext™ 5-mC RNA Bisulfite-Seq Easy Kit (Illumina).

產品列表

貨號	產品	數量
P-9007-12	EpiNext 5-mC RNA Bisulfite-Seq Easy Kit (Illumina) 5-mc RNA亞硫酸鹽-定序簡易試劑套組 (組) 廠牌:Epigentek	12 reactions
P-9007-24	EpiNext 5-mC RNA Bisulfite-Seq Easy Kit (Illumina) 5-mc RNA亞硫酸鹽-定序簡易試劑套組 (組) 廠牌:Epigentek	24 reactions

EpiNext 5-mC RNA Bisulfite-Seq Easy Kit (Illumina)

• High sensitivity and efficiency
• Easy and streamlined procedure
• Complete conversion (>99.9%)
• Extremely convenient
• Minimal bias
• Broad sample suitability