whole genome sequencing with steps.

1. Whole Genome Sequencing
By: Qadardana kakar
BS Biotechnology
7th semester

2. Aims/Objective
 The aim of my presentation is to elucidate the Whole Genome Sequencing
technique and its protocol.

3. Discovery
 Sanger and his colleagues invented chain termination sequencing technique in 1970.
 Fragments of 100-1000 base pair.
 In 1979 shotgun sequencing technique was developed.
 Large size fragments.
 Overlapping of discovered fragments to detect whole genome sequences.

4. WGS Whole Genome Shotgun Sequencing
 To sequence all genome of a particular organism.
1. Find sequence.
2. Find their position in entire genome.
steps
 Collect and isolate the DNA or genome.
 Break into small manageable pieces.
 Copy each piece many times.
 Read the DNA sequence
 Assemble the data into a genome.

5. WGS
1. Different techniques are used to cut DNA into particular size pieces.
 Fragments are electrophoresed through gel to find the size of fragments.
2. Ligation of fragment into a plasmid.

6. 4. Insert plasmid into bacteria e.g. E.coli.
Bacteria with plasmid select Transformed bacteria (incubated)

7.  5. Microtiter plate containing E.coli is heated to 95 degree Celsius.
 plasmids are released.
 6. PCR reaction amplify the plasmid.
Rolling circle amplification

8.  7. Desired sequence amplification (Stephenson, 2003).

9. 8. DNA attracted toward the carboxyl coated magnetic bead.
 In Strong ethanol solution the carboxyl coat attracts the phosphate backbone.
 Cellular debris and reagents remain in solution.
 Wash with ethanol. Water added to each well.
 Water causes DNA to be released.

10. 9.Sanger sequencing.
 10. Assembly
 Multiple read of the same genome is made.
 Reassembles by matching overlaps.

11. All Pictures are retrieved from: Whole genome sequencing/ YouTube.

12. Whole Genome Sequencing
Advantages
 To find coding non and non coding regions.
 Personalized drugs.
 Disease susceptibility prediction.
 Specie comparison and evolutionary studies.