1. CS262 Discussion Section 3

2. Topics for today DNA replication DNA sequencing: Biological tools Transposons “Out of Africa” hypothesis of human origins

3. At each cell division, a cell must copy its genome with extraordinary accuracy

4. Base pairing enables DNA replication Designate the two DNA strands as S and S’ Strand S separates from strand S’. Strand S can serve as a template for making a new strand S’, while strand S’ can serve as a template for making a new strand S. http://www.contexo.info/DNA_Basics/images/ DNAreplication.mov http://www.contexo.info/DNA_Basics/images/ DNAreplication.mov

5. DNA synthesis begins at replication origins Replication origin: The position at which the DNA is first opened. DNA polymerase cannot start a completely new DNA strand, it can join a new nucleotide only to a base-paired nucleotide in the double helix. DNA polymerase can elongate a new DNA chain only in the 5’-3’ direction.

6. Short lengths of RNA act as primers for DNA synthesis A different enzyme is needed to begin a new DNA strand – primase. A short length of RNA (~10 nucleotides) serves as a primer for DNA synthesis. Show animation from Lodish et al.Lodish et al

7. DNA replication is semi-conservative

8. Physically sorting DNA molecules according to their lengths

9. Gel electrophoresis sorts DNA fragments of different lengths 1. Mixture of DNA fragments loaded at one end of a gel, which contains a microscopic network of pores. 2. A voltage is applied across the gel slab. 3. The fragments migrate towards the +ve electrode, because DNA is –vely charged. 4. Larger fragments migrate more slowly because their progress is more impeded by the gel matrix.

10. Gel electrophoresis sorts DNA fragments of different lengths 5. The DNA fragments become spread out across the gel according to size (length), forming a ladder of discrete bands. Each band is composed of a collection of DNA molecules of identical length. 6. The bands are invisible unless the DNA is labeled in some way. 7. A radioisotope is incorporated into the DNA molecules before electrophoresis, eg 32 P

11. Gel electrophoresis sorts DNA fragments of different lengths 8. Because the particles emitted from 32 P will expose photographic film, a sheet of film placed flat on top of the gel will (when developed) show the position of all the DNA bands.

13. DNA Sequencing Phosphate Group Sugar Nitrogenous Base

15. ANIMATION on SANGER’S DIDEOXY METHOD

16. The mystery of the primer How do we know what the primer is?? Hint: Recall restriction enzymes.

17. Some DNA can jump Transposition is the movement of genetic material from one chromosomal location (donor site) to another (target site). DNA sequences that possess this capability to change their genomic location are called mobile elements or transposable elements. There are two types of transposition, depending on whether the transposable element is replicated or not.

18. 1. Conservative transposition (~Cut-and-paste) The element itself moves from one site to another. Transposable Element Target RecipientDonor Excision & Integration

19. 2. Replicative transposition (~Copy-and-paste) The transposable element is copied, and one copy remains at the original site while the other inserts itself at a new site. Transposable Element Target RecipientDonor Integration Replication

20. Transposons in bacteria They are mobile elements that carry the transposase gene (the enzyme that disconnects the transposon from the flanking DNA and inserts into the target DNA site) and may also carry genes that inactivate antibiotics like ampicillin and tetracycline.

21. Retrotransposons 1. DNA of the mobile element is transcribed into RNA. 2. This RNA is reverse-transcribed into cDNA, which then inserts into the recipient site. Target Donor Integration Transcription RNA cDNA Reverse Transcription

22. Alu sequence 300 nucleotide pairs Present in about 1 million copies in the human genome. Constitutes 11% of human DNA. Do not encode their own reverse transcriptase, depend on enzymes already present in the cells to help them move.

23. What is the origin of our species?

24. Africa – the cradle of humans? Notable events in human evolution are said to be the gradual abandonment of tree- dwelling existence and the switch to bipedalism (two-legged walking). All human evolution took place in Africa, more specifically in southern and eastern Africa.

25. ‘Out of Africa to your hometown’ or Multi-regionalism? http://www.becominghuman.org

26. Migration of human variation http://info.med.yale.edu/genetics/kkidd/point.html

27. Migration of human variation http://info.med.yale.edu/genetics/kkidd/point.html

28. Migration of human variation http://info.med.yale.edu/genetics/kkidd/point.html