Next Generation Sequencing Overview (recording)

These flashcards cover the key concepts related to next generation sequencing techniques, including workflows, types, gene expression, and analysis methods.

What is next generation sequencing often referred to as?

High throughput sequencing technologies.

What is a DNA library?

A collection of DNA fragments that collectively represent the entire genome.

How does high throughput sequencing differ from traditional Sanger sequencing?

High throughput sequencing bypasses the cloning step and allows for massive parallel sequencing of DNA fragments.

What does 'massive parallel sequencing' mean?

It refers to sequencing many fragments simultaneously.

What is a reference sequence?

A sequence used as a guide to assemble other sequences by aligning them in the correct order.

What happens during the sequencing by synthesis process?

DNA is sequenced as nucleotides are added, with the sequence being read by a detector.

What distinguishes second generation sequencing from third generation sequencing?

Second generation sequencing requires amplification of fragments before sequencing, while third generation sequencing does not.

What is the role of the coverage in sequencing?

Coverage indicates the average number of reads that align to known reference bases, determining the certainty of variant discovery.

What is nanopore sequencing?

A sequencing method that detects differences in electrical conductivity as single-stranded DNA passes through a nanopore.

What is the primary focus of gene expression analysis?

To determine which genes are expressed in different cells or conditions, contributing to their distinct phenotypes.

What is a Northern Blot used for?

To detect specific RNA molecules through gel electrophoresis and hybridization with labeled DNA probes.

What does the term 'transiently expressed' refer to in gene expression studies?

A gene that shows expression only for a short period in response to stimuli.

What mechanism do researchers use to ensure consistent RNA levels in a gel analysis?

Using a housekeeping gene as an internal control to compare RNA amounts.

What is the main purpose of a Western Blot?

To detect specific proteins in a sample after separation by SDS-PAGE.

What does the term 'depth of coverage' indicate in sequencing?

The number of times a specific position in the genome is sequenced.

Why is it important to use both experimental and control samples in gene expression studies?

To differentiate between actual changes in gene expression and variations due to different RNA levels.