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Quality evaluation of Methyl Binding Domain based kits for enrichment DNA-methylation sequencing

ABSTRACT: Purpose: to compare different Methyl Binding Domain (MBD) based kits for DNA-methylation sequencing using Reduced Representation Bisulfite Sequencing (RRBS) data for validation, and to determine whether data quality can also be derived from inherent sequence data characteristics

ORGANISM(S): Homo sapiens

PROVIDER: GSE42790 | GEO | 2013/01/23

SECONDARY ACCESSION(S): PRJNA183137

REPOSITORIES: GEO

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Quality evaluation of Methyl Binding Domain based kits for enrichment DNA-methylation sequencing

Project description:Purpose: to compare different Methyl Binding Domain (MBD) based kits for DNA-methylation sequencing using Reduced Representation Bisulfite Sequencing (RRBS) data for validation, and to determine whether data quality can also be derived from inherent sequence data characteristics MBD-seq using 5 different kits (MethylCap, MethylCollector, MethylCollector Ultra, MethylMiner, MethylMagnet) was applied on 3 commonly used cell lines (DU145, HCT15, PC3), for which also RRBS data were generated.

2013-01-23 | E-GEOD-42790 | biostudies-arrayexpress

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Quality evaluation of methyl binding domain based kits for enrichment DNA-methylation sequencing.

Project description:DNA-methylation is an important epigenetic feature in health and disease. Methylated sequence capturing by Methyl Binding Domain (MBD) based enrichment followed by second-generation sequencing provides the best combination of sensitivity and cost-efficiency for genome-wide DNA-methylation profiling. However, existing implementations are numerous, and quality control and optimization require expensive external validation. Therefore, this study has two aims: 1) to identify a best performing kit for MBD-based enrichment using independent validation data, and 2) to evaluate whether quality evaluation can also be performed solely based on the characteristics of the generated sequences. Five commercially available kits for MBD enrichment were combined with Illumina GAIIx sequencing for three cell lines (HCT15, DU145, PC3). Reduced representation bisulfite sequencing data (all three cell lines) and publicly available Illumina Infinium BeadChip data (DU145 and PC3) were used for benchmarking. Consistent large-scale differences in yield, sensitivity and specificity between the different kits could be identified, with Diagenode's MethylCap kit as overall best performing kit under the tested conditions. This kit could also be identified with the Fragment CpG-plot, which summarizes the CpG content of the captured fragments, implying that the latter can be used as a tool to monitor data quality. In conclusion, there are major quality differences between kits for MBD-based capturing of methylated DNA, with the MethylCap kit performing best under the used settings. The Fragment CpG-plot is able to monitor data quality based on inherent sequence data characteristics, and is therefore a cost-efficient tool for experimental optimization, but also to monitor quality throughout routine applications.

| S-EPMC3598902 | biostudies-literature

Evaluation of methyl-binding domain based enrichment approaches revisited

Project description:A comparison of methylMiner and MethylCap, two methylation enrichment kits The main analysis include two individuals (mice) in technical duplicates from various lab/technical conditions. The results are compared with previous studies of two human samples from a number of lab/technical conditions (human data available at dbGaP: record number phs000608.v1.p1). In addition a DNA from a third mouse is used for investigations of specific alterations of the protocol.

2014-12-20 | E-GEOD-64144 | biostudies-arrayexpress

Evaluation of three small RNA library preparation kits on serum-derived RNA

Project description:We evaluated the effect of the small RNA library preparation method on 5' tRNA-halves and miRNA abundance in libraries prepared from serum RNA using three commercially available small RNA library preparation kits (TruSeq small RNA library preparation kit v2 (Illumina), TailorMix miRNA sample preparation kit v2 (Seqmatic) and the NEBNext Multiplex Small RNA library prep kit (New England Biolabs)). RNA isolated from 100 µl of serum collected from healthy mice was used as input for the preparation of a small RNA library in duplicate and libraries were single end sequenced.

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Systematic assessment of commercially available low-input miRNA library preparation kits

Project description:We compared the performance of six commercial kits (QIAseq, SMARTer, CATS, CleanTag, srLp, TailorMix) capable of handling <100ng total RNA input for miRNA detection sensitivity, reliability, titration response and ability to detect differentially expressed miRNAs at different amounts of synthetic miRNAs. We observed differences in detection sensitivity between the kits, but none were able to detect the full repertoire of expected miRNAs. The reliability within the replicates of all kits was good, while larger differences were observed between the kits, although none could accurately quantify the majority of miRNAs.

2019-09-27 | GSE133719 | GEO

Quality evaluation of Enzymatic Methyl-seq library constructed using crude cell lysate

Project description:Enzymatic Methyl-seq (EM-seq) is an enzyme-based method giving us reliable methylome data from small amounts of purified DNA. However, it is unclear whether EM-seq library can be constructed from crude cell lysate containing genomic DNA. We evaluated the quality of EM-seq libraries directly prepared from crude cell lysate.

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An extension to: Systematic assessment of commercially available low-input miRNA library preparation kits

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2020-05-31 | GSE141658 | GEO

Profiling DNA Methylation Differences Between Inbred Mouse Strains on the Illumina Human Infinium MethylationEPIC Microarray

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Cross-Site Comparison of Ribosomal Depletion Kits for Illumina RNAseq Library Construction

Project description:In a cross-site study we evaluated the performance of ribosomal RNA removal kits from Illumina, Takara/Clontech, Kapa Biosystems, Lexogen, New England Biolabs and Qiagen on intact and degraded RNA samples. We found that all of the kits were capable of performing significant ribosomal depletion, though there were differences in their ease of use. All kits were able to remove ribosomal RNA to below 20% with intact RNA and identify ~14,000 protein coding genes from the Universal Human Reference RNA sample at >1FPKM. Analysis of differentially detected genes among kits suggested that transcript length may be a key factor in library production efficiency. These results provide a roadmap for labs on the strengths of each of these methods and how best to utilize them.

2018-02-01 | GSE100127 | GEO

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