Strand-specific Sequencing
Strand-specific Sequencing - Related Videos
08:53
Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method
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2025
Enrichment and sequencing of protein-associated nascent DNA (eSPAN) was developed to detect the relative abundance of a chromatin-associated protein on two replicating DNA strands, thereby revealing molecular insight into chromatin replication and its coupled processes. This protocol describes eSPAN procedures in yeast and mouse embryonic stem (ES) cells.
09:04
Sequence-specific and Selective Recognition of Double-stranded RNAs over Single-stranded RNAs by Chemically Modified Peptide Nucleic Acids
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Cited by 12 •
2017
We report the protocols for the synthesis and purification of Peptide Nucleic Acid (PNA) oligomers incorporating modified residues. The biochemical and biophysical methods for the characterization of the recognition of RNA duplexes by the modified PNAs are described.
03:20
Nested-PCR to Detect a Specific Viral Genomic Sequence
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2025
This video describes nested polymerase chain reaction, a technique that consists of two sequential PCR amplification processes using two primer sets. The first set of primers is intended to anneal to sequences upstream of the second set, resulting in selective amplification of specific gene sequences. This PCR is more sensitive and specific than a normal PCR and is widely used as a detection technique for various diseases.
12:12
Analysis of Termination of Transcription Using BrUTP-strand-specific Transcription Run-on (TRO) Approach
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Cited by 10 •
2017
We describe a basic experimental approach for analysis of termination of transcription by RNA polymerase II in vivo using BrUTP by the strand-specific transcription run-on (TRO) approach in budding yeast. This protocol can be extended to study transcription termination by other RNA polymerases both in yeast and higher eukaryotes.
02:04
Fixing Double-strand Breaks
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2020
The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
FAQs
During library preparation, directional adapters or strand-selective reverse transcription preserve information about the source strand before sequencing. Related directional workflows maintain that signal through construction of the sequencing library. Afterward, reads are aligned according to their orientation, linking observed sequences to the appropriate sense or antisense transcript rather than treating strand identity as ambiguous.
Orientation is especially informative when transcripts overlap or when a locus produces antisense RNA. Without strand information, reads from opposite transcriptional directions can be difficult to assign to the correct feature. Retaining orientation therefore helps separate overlapping genes, identify antisense transcription, and improve transcript annotation and abundance estimates in complex transcriptomes.
Compared with conventional sequencing, Strand-specific sequencing adds directional evidence to read interpretation. That distinction can reveal whether signal belongs to a sense or antisense transcript, rather than combining contributions from both orientations. The result is more informative analysis of transcript abundance, alternative transcription, and noncoding RNA expression when multiple transcript features occupy the same genomic region.
A typical workflow begins by preparing a directional library using adapters, strand-selective reverse transcription, or a related strand-preserving strategy. The resulting material is sequenced, and reads are aligned to a reference genome or transcriptome with orientation retained. Researchers can then use the directional alignments to annotate transcripts and evaluate expression patterns that depend on strand assignment.
It is most useful when a sample contains overlapping genes, antisense RNA, or a complex mixture of coding and noncoding transcripts. In these settings, strand-aware measurements help researchers distinguish transcriptional sources and characterize gene regulation or genome organization. The approach also supports studies of alternative transcription and transcript abundance, where assigning reads to the correct orientation affects interpretation.
They can clarify which transcriptional direction contributes to a read signal and thereby improve interpretation of transcript features. In transcriptome analysis, this information supports more precise annotation, separates sense from antisense expression, and helps assess noncoding RNA activity. In biology, those outcomes connect sequencing measurements with questions about gene regulation, genome organization, and how complex transcriptomes are arranged.