Question

I Q10A. Differentiate between two types of DNA sequencing methodologies, sequencing by chain termination and next generation sequencing. (5 points)

Answer 1

In the year 1977, Fredrick Sanger postulated the first method for sequencing the DNA, named as a chain termination method. The chain termination method is also referred to as a dideoxynucleotide sequencing because of the use of the special types of ddNTPs. The ddNTPs are different from normal dNTPs. it possesses the hydrogen group instead of a hydroxyl group in the dNTPs.

The process of Sanger sequencing is broadly divided into 3 steps:

1. DNA extraction: using any of the DNA extraction protocols
2. PCR amplification: using the flanking primers, dNTPs, ddNTPs, Taq DNA polymerase and PCR buffer.
3. Identification of the amplified fragments: using autoradiography, PAGE, or capillary gel electrophoresis.

The older, classical chain termination method is also called the Sanger method. Newer methods that can process a large number of DNA molecules quickly are collectively called High-Throughput Sequencing (HTS) techniques or Next-Generation Sequencing (NGS) methods.

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