DNA Sequencing

What is DNA Sequencing?

• Is the process of determining the
sequence of nucleotide bases (As, Ts,
Cs, and Gs) in a piece of DNA.
• Is the determination of the precise
sequence of nucleotides in a sample
of DNA.
• Is technologies have gone through at
least three "generations": Sanger
sequencing and Gilbert sequencing
were first-generation,
pyrosequencing was second-
generation, and Illumina sequencing
is next-generation.
 Benchmark of DNA sequencing

• 1953 Discovery of the structure of the DNA double helix. 

• 1972 Development of recombinant DNA technology, which permits isolation of defined fragments of DNA; prior to this,
the only accessible samples for sequencing were from bacteriophage or virus DNA. 
• 1975 The first complete DNA genome to be sequenced is that of bacteriophage φX174 
• 1977 Allan Maxam and Walter Gilbert publish "DNA sequencing by chemical degradation". Fred Sanger, independently,
publishes "DNA sequencing by enzymatic synthesis".
• 1990 The U.S. National Institutes of Health (NIH) begins large-scale sequencing trials on Mycoplasma capricolum,
Escherichia coli, Caenorhabditis elegans, and Saccharomyces cerevisiae (at 75 cents (US)/base).
• 2001 HGP consortium publishes Human Genome Sequence draft in Nature (15 Feb).  Celera publishes the Human
Genome sequence. 
• 2005 420,000 VariantSEQr human resequencing primer sequences published on new NCBI Probe database. 
• 2007 For the first time, a set of closely related species (12 Drosophilidae) are sequenced, launching the era of
phylogenomics.  Craig Venter publishes his full diploid genome: the first human genome to be sequenced completely.
• 2008 An international consortium launches The 1000 Genomes Project, aimed to study human genetic variability.
• 2008 Leiden University Medical Center scientists decipher the first complete DNA sequence of a woman.
 Methods in DNA Sequencing

The general method of DNA sequencing is as follow:

1. The piece of chromosomes is taken must contains 50 to 250 million nucleotide bases. These bases are then
converted into smaller pieces.
2. The pieces are used as templates to produce set of fragments of different length.
3. Used agarose DNA gel electrophoresis to separate the fragments and keep them in order.
4. Fluorescent dyes are used to separate the set of fragments on the agarose gel and keep them in sequence.
5. The final bases, at the end of each strand/fragment are identified which when combined together make the
short strand which was generated in the first set.
6. Automated sequencers are used for analysis of resulting electropherograms, they show four color
chromatogram in which peaks are showing these peaks represent the four DNA bases.
7. After the visualization of the bases, now computers are use and all the four fragments are combined together
to make a long stretch. This long fragments are analyzed for gene coding regions, errors, and for many other
characteristics.
 Benefits, Application and Risk of DNA sequencing
Benefits:
• Enables the scientists to determines genome sequence of living organisms.
• It helps the scientists to identify genes which are responsible for causing genetic diseases and disorder.

Applications:
• Forensics – DNA sequencing applied in forensics science to identify particular individual because every individual has
unique sequence of its genetic data. It is particularly used to identify the criminals in finding proof from crime scene in the
form of hair, nail, skin or blood samples.
• Medicine – in medical research, DNA sequencing can be used to detect the genes which are associated with some heredity
or it acquired disease.
• Agriculture – DNA sequence played a vital role in the field of agriculture. It is used in mapping and sequencing the whole
genome of microorganisms to make them useful. Example, specific genes of bacteria have been used to increase their
resistance to insects and pests to increase the productivity and nutritional value of the plant. Similarly, it has been useful to
livestock production with improved quality of milk and meat.

Risks:
• infer paternity or other features of the participant's genealogy
• claim statistical evidence that could affect employment or insurance or the ability to obtain financial services for the
participant
• claim relatedness to criminals or incriminate relatives
• make synthetic DNA corresponding to the participant and plant it at a crime scene
• reveal propensity for a disease currently lacking effective treatment options
7 Approaches used in DNA sequencing