Other methods address different types of DNA modifications using the MinION platform. Stoiber et al. examined 4-methylcytosine (4mC) and 6-methyladenine (6mA), along with 5mC, and also created a software to directly visualize the raw MinION data in  human-friendly way. Here they found that in E. coli, which has a known methylome, event windows of 5 base pairs long can be used to divide and statistically analyze the raw MinION electrical signals. A straightforward Mann-Whitney U test can detect modified portions of the E. coli sequence, as well as further split the modifications into 4mC, 6mA or 5mC regions. PacBio sequencing has also been used to detect DNA methylation. In this platform  the pulse width - the width of a fluorescent light pulse - corresponds to a specific base. In 2010 it was shown that the interpulse distance in control and methylated samples are different, and there is a "signature" pulse width for each methylation type. In 2012 using the PacBio platform the binding sites of DNA methyltransferases were characterized. The detection of N6-methylation in C Elegans was shown in 2015. DNA methylation on N6-adenine using the PacBio platform in mouse embryonic stem cells was shown in 2016. While expression levels as the gene level can be more or less accurately depicted by second generation sequencing, transcript level information is still an important challenge. As a consequence, the role of alternative splicing in molecular biology remains largely elusive. Third generation sequencing technologies hold promising prospects in resolving this issue by enabling sequencing of mRNA molecules at their full lengths. A study published in 2008 surveyed 25 different existing transcript reconstruction protocols. Its evidence suggested that existing methods are generally weak in assembling transcripts, though the ability to detect individual exons are relatively intact. According to the estimates, average sensitivity to detect exons across the 25 protocols is 80\% for Caenorhabditis elegans genes. In comparison, transcript identification sensitivity decreases to 65\%. For human, the study reported an exon detection sensitivity averaging to 69\% and transcript detection sensitivity had an average of  mere 33\%. In other words, for human, existing methods are able to identify less than half of all existing transcript.
Other methods address different types of DNA modifications using the MinION platform. Stoiber et al. examined 4-methylcytosine (4mC) and 6-methyladenine (6mA), along with 5mC, and also created software to directly visualize the raw MinION data in a human-friendly way. Here they found that in E. coli, which has a known methylome, event windows of 5 base pairs long can be used to divide and statistically analyze the raw MinION electrical signals. A straightforward Mann-Whitney U test can detect modified portions of the E. coli sequence, as well as further split the modifications into 4mC, 6mA or 5mC regions. PacBio sequencing has also been used to detect DNA methylation. In this platform , the pulse width - the width of a fluorescent light pulse - corresponds to a specific base. In 2010 it was shown that the interpulse distance in control and methylated samples are different, and there is a "signature" pulse width for each methylation type. In 2012 using the PacBio platform the binding sites of DNA methyltransferases were characterized. The detection of N6-methylation in C Elegans was shown in 2015. DNA methylation on N6-adenine using the PacBio platform in mouse embryonic stem cells was shown in 2016. While expression levels as the gene level can be more or less accurately depicted by second generation sequencing, transcript-level information still remains an important challenge. As a consequence, the role of alternative splicing in molecular biology remains largely elusive. Third generation sequencing technologies hold promising prospects in resolving this issue by enabling sequencing of mRNA molecules at their full lengths. A study published in 2008 surveyed 25 different existing transcript reconstruction protocols. Its evidence suggested that existing methods are generally weak in assembling transcripts, though the ability to detect individual exons are relatively intact. According to the estimates, average sensitivity to detect exons across the 25 protocols is 80\% for Caenorhabditis elegans genes. In comparison, transcript identification sensitivity decreases to 65\%. For human, the study reported an exon detection sensitivity averaging to 69\% and transcript detection sensitivity had an average of a mere 33\%. In other words, for human, existing methods are able to identify less than half of all existing transcript.
D
a
A
a
A
,
R
transcript level information is still
transcript-level information still remains
A
a