____________ is the process of determining the order of nucleotides in a DNA molecule.
DNA sequencing
What are the 4 major Nucleotides?
- Adenine (A)
- Thymine (T)
- Cytosine (C)
- Guanine (G)
The order of ___________ in a DNA molecule determines the ___________
nucleotides; genetic code
_____________ the set of instructions for building and maintaining an organism.
genetic code
What are the two main types of DNA sequencing methods?
- Sanger method
- next-generation sequencing (NGS)
chain-termination method
Sanger sequencing
Sanger sequencing uses _____________ to terminate DNA synthesis.
dideoxynucleotides (ddNTPs)
Dideoxynucleotides are similar to nucleotides, but they lack a ____________ which is necessary for DNA synthesis to continue
3’-hydroxyl group,
To perform Sanger sequencing, a DNA sample is amplified using ________ and then incubated with a mixture of ________________ The ddNTPs will randomly terminate DNA synthesis, resulting in a series of DNA fragments of different lengths. The DNA fragments are then separated by ______________ and the sequence of the DNA molecule is determined by reading the order of the bands on the gel.
- PCR
- ddNTPs and dNTPs
- gel electrophoresis
_______________ is a collection of methods that can sequence DNA much faster and more cheaply than Sanger sequencing.
Next-generation sequencing
What are the advantages of NGS?
sequence much faster & more cheap then Sanger sequencing
NGS methods typically involve __________________, meaning that many DNA molecules are sequenced simultaneously
massively parallelizing the sequencing process
Two techniques employed in NGS?
- Sequencing-by-synthesis (SBS)
- Sequencing-by-ligation (SBL)
NGS methods have revolutionized the _____________ making it possible to sequence entire genomes at a relatively low cos
field of genomics
What are examples of wide applications of NGS?
- research,
- diagnostics,
- and personalized medicine
the first DNA sequencing method to be developed
Sanger sequencing
Give the advantages and disadvantages of Sanger sequencing
Pros:
Relatively simple and Straightforward
Cons:
Time consuming and expensive to sequence large DNA molecules
What are the steps involved in Sanger sequencing?
- The DNA sample is amplified using PCR.
- The amplified DNA is incubated with a mixture of ddNTPs and dNTPs.
- The ddNTPs will randomly terminate DNA synthesis, resulting in a series of DNA fragments of different lengths.
- The DNA fragments are separated by gel electrophoresis.
- The order of the nucleotides in the DNA molecule is determined by reading the order of the bands on the gel.
the most common type of NGS method
Sequencing-by-synthesis
how does SBS work?
It works by sequencing the DNA molecule one nucleotide at a time.
used to image the surface and determine the order of the nucleotides in the DNA molecule.
laser
how does SBL work?
sequencing the DNA molecule one fragment at a time
contain sequences that are known to bind to the sequencing machine
adapters
reads the order of the nucleotides in the chain.
Sequencing machine
