Sentence examples for non-converted DNA from inspiring English sources
Pyrosequence analysis of bisulfite converted and non-converted DNA was performed by EpigenDx (Worcester, MA, USA).
Hence, we optimised the melting temperature with gradient PCR, using both converted and non-converted DNA and water to minimise false-positives and these controls were included in every amplification step from then on.
We analysed the specificity of the mir-124-3 mir-124-3 mir-124-3plicate measurements of converted methylated (M), converted non-methylated (U) and non-converted DNA control samples.
The analysis of converted methylated (M), converted non-methylated (U), and non-converted DNA control samples demonstrated that the NEFH qMSP specifically detects M DNA while U and non-converted samples remained undeterminable both in the QC1 control reaction as well as in the NEFH-specific PCR (Ct > 45 cycles, Figure 1B).
The non-converted DNA did not give rise to amplifications for both primer sets.
Semi-nested regular PCR was chosen to minimise the risk of amplifying non-converted DNA.
Bisulphite-converted methylated and unmethylated human DNA, as well as non-converted unmethylated DNA that was converted along with the samples, and pure water, were included with each PCR analysis as positive or negative controls.
Given that the total number of usable reads does not change substantially (even when the proportion of a lane comprising bisulfite-converted libraries is decreased by 50%) with increasing proportion of non-bisulfite converted DNA, we would suggest that bisulfite converted libraries can be run with non-bisulfite converted libraries at minimal cost to sequencing output.
In this work we carefully and completely lay out experimental procedures and QC parameters to assess BisSeq experiments and we suggest that pooling 30% non-bisulfite converted DNA with bisulfite converted DNA has a minimal effect on bisulfite read output, meaning that sequencing non-bisulfite DNA samples from unrelated experiments is practical.
PCR was performed with primers annealing to the non bisulfite-converted DNA (5′-CCAACCTGACTGTGGTGGACAA-3; 5′-ACATGCACCTTCCCAGGGC-3′).
We also determine that including a significant portion of non-bisulfite converted DNA with bisulfite converted DNA has a minimal impact on usable bisulfite read output.
