Limits...
Design, measurement and processing of region-specific DNA methylation assays: the mass spectrometry-based method EpiTYPER.

Suchiman HE, Slieker RC, Kremer D, Slagboom PE, Heijmans BT, Tobi EW - Front Genet (2015)

Bottom Line: The technology targets genomic regions of 100-600 base pairs and results in the quantitative measurement of DNA methylation levels largely at single-nucleotide resolution.It is particularly suitable for larger scale efforts to study candidate regions or to validate regions from genome-wide DNA methylation studies.Here, we describe in detail how to design and perform EpiTYPER measurements and preprocess the data, providing details for high quality measurements not provided in the standard EpiTYPER protocol.

View Article: PubMed Central - PubMed

Affiliation: Molecular Epidemiology, Department of Medical Statistics and Bioinformatics, Leiden University Medical Center Leiden, Netherlands.

ABSTRACT
EpiTYPER® is a mass spectrometry-based bisulfite sequencing method that enables region-specific DNA methylation analysis in a quantitative and high-throughput fashion. The technology targets genomic regions of 100-600 base pairs and results in the quantitative measurement of DNA methylation levels largely at single-nucleotide resolution. It is particularly suitable for larger scale efforts to study candidate regions or to validate regions from genome-wide DNA methylation studies. Here, we describe in detail how to design and perform EpiTYPER measurements and preprocess the data, providing details for high quality measurements not provided in the standard EpiTYPER protocol.

No MeSH data available.


Example of mass spectra resulting from the measurement with EpiTYPER. (A) Shows an entire mass spectrum of a single measurement of a sample. Black peaks are measured fragments while the blue and red dotted lines denote a selected CpG fragment in the EpiTYPER software. This particular part of the mass spectrum is shown in (B). The spectrum is nice and smooth, indicating a good quality of the measurement.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4585020&req=5

Figure 5: Example of mass spectra resulting from the measurement with EpiTYPER. (A) Shows an entire mass spectrum of a single measurement of a sample. Black peaks are measured fragments while the blue and red dotted lines denote a selected CpG fragment in the EpiTYPER software. This particular part of the mass spectrum is shown in (B). The spectrum is nice and smooth, indicating a good quality of the measurement.

Mentions: First, we inspected at the entire mass spectrum of several of these 12 triplicate measurements. The measurements all showed a nice baseline following the x-axis (Figure 5A), indicative of little measurement noise, and the signals of the predicted peaks are high. Moreover, additional peaks were distributed randomly across the measurements and lower in intensity than the predicted peaks. Together, this is indicative for optimal spotting and laser intensity settings, little unspecific cleavage and no non-specific PCR product formation (see Note 5). When looking at some CpG units in more detail the spectra were smooth (Figure 5B), again indicative of a good quality measurement.


Design, measurement and processing of region-specific DNA methylation assays: the mass spectrometry-based method EpiTYPER.

Suchiman HE, Slieker RC, Kremer D, Slagboom PE, Heijmans BT, Tobi EW - Front Genet (2015)

Example of mass spectra resulting from the measurement with EpiTYPER. (A) Shows an entire mass spectrum of a single measurement of a sample. Black peaks are measured fragments while the blue and red dotted lines denote a selected CpG fragment in the EpiTYPER software. This particular part of the mass spectrum is shown in (B). The spectrum is nice and smooth, indicating a good quality of the measurement.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4585020&req=5

Figure 5: Example of mass spectra resulting from the measurement with EpiTYPER. (A) Shows an entire mass spectrum of a single measurement of a sample. Black peaks are measured fragments while the blue and red dotted lines denote a selected CpG fragment in the EpiTYPER software. This particular part of the mass spectrum is shown in (B). The spectrum is nice and smooth, indicating a good quality of the measurement.
Mentions: First, we inspected at the entire mass spectrum of several of these 12 triplicate measurements. The measurements all showed a nice baseline following the x-axis (Figure 5A), indicative of little measurement noise, and the signals of the predicted peaks are high. Moreover, additional peaks were distributed randomly across the measurements and lower in intensity than the predicted peaks. Together, this is indicative for optimal spotting and laser intensity settings, little unspecific cleavage and no non-specific PCR product formation (see Note 5). When looking at some CpG units in more detail the spectra were smooth (Figure 5B), again indicative of a good quality measurement.

Bottom Line: The technology targets genomic regions of 100-600 base pairs and results in the quantitative measurement of DNA methylation levels largely at single-nucleotide resolution.It is particularly suitable for larger scale efforts to study candidate regions or to validate regions from genome-wide DNA methylation studies.Here, we describe in detail how to design and perform EpiTYPER measurements and preprocess the data, providing details for high quality measurements not provided in the standard EpiTYPER protocol.

View Article: PubMed Central - PubMed

Affiliation: Molecular Epidemiology, Department of Medical Statistics and Bioinformatics, Leiden University Medical Center Leiden, Netherlands.

ABSTRACT
EpiTYPER® is a mass spectrometry-based bisulfite sequencing method that enables region-specific DNA methylation analysis in a quantitative and high-throughput fashion. The technology targets genomic regions of 100-600 base pairs and results in the quantitative measurement of DNA methylation levels largely at single-nucleotide resolution. It is particularly suitable for larger scale efforts to study candidate regions or to validate regions from genome-wide DNA methylation studies. Here, we describe in detail how to design and perform EpiTYPER measurements and preprocess the data, providing details for high quality measurements not provided in the standard EpiTYPER protocol.

No MeSH data available.