lecture 7 - sequencing Flashcards
(22 cards)
Who developed Sanger sequencing and in which decade?
Fred Sanger in the 1970s.
What method was originally used in Sanger sequencing?
Radiolabeling.
What are modified bases used in Sanger sequencing?
ddNTPs (dideoxynucleotides).
What does Sanger sequencing end with that allows reading the DNA sequence?
A ladder that can be read upwards.
What technology replaced radioactivity in Sanger sequencing?
Fluorescence.
What types of mutations can Sanger sequencing identify?
Point and indel mutations.
What is the average size of DNA fragments used in Next-Generation Sequencing (NGS)?
Approximately 800 bp.
What device is used to shear DNA samples in NGS?
A nebulizer.
What are the size ranges of fragments isolated in NGS?
150-200 bp.
What sequencing method does NGS rely on?
Polony sequencing.
What are the typical read lengths in NGS?
50-250 bp.
What is the range of read lengths in Sanger sequencing?
1-2 kbp (1000-2000 bp).
What is required for Sanger sequencing that is not necessary for NGS?
Knowledge of what you are sequencing.
What does ChIPseq stand for?
Chromatin immunoprecipitation sequencing.
What mutation is highlighted in the chromatogram example involving CLDN19?
A nucleotide change C to T at chr1:43,205,676.
What does the chromatogram show for the index patient at chr1:43,205,676?
Homozygous T/T.
What type of mutation is represented by the change c.1061C> T?
A missense mutation.
In the chromatogram example, what does a double peak represent?
A heterozygous genotype.
What does a single peak in a chromatogram indicate?
A homozygous genotype.
What mutation is illustrated by the deletion of two nucleotides AG in the BRCA1 gene?
85delAG.
What is a key advantage of DNA sequencing for diagnosing genetic disorders?
It offers key advantages over DNA hybridization and PCR-based techniques.
How has DNA sequencing technology evolved?
To provide reads over a larger region of DNA in a shorter amount of time.
