1. Chapter 20 DNA Technology and Genomics

2. Chapter 20 DNA Technology and Genomics
DNA cloning
DNA probe hybridization
Polymerase Chain Reaction (PCR)
Gel Electrophoresis
Southern Blot
DNA sequencing.

3. Chapter 20 DNA Cloning
Bacteria have restriction enzymes to attack and destroy invading viral DNA. Restriction enzymes cut DNA at specific nucleotide sequences leaving “sticky ends.” DNA ligase can seal these ends, making recombinant DNA.

4. Chapter 20 DNA Cloning
Restriction fragments can be put into plasmids. Gene cloning occurs when cells containing these plasmids reproduce. Genes of interest are marked with a radioactive DNA probe.

5. Chapter 20 DNA Cloning
If a gene is inserted next to a promoter, the bacteria becomes an expression vector. Eukaryotic chromosomes allow for bigger segments of DNA. Eukaryotic cells can also process polypeptides into proteins.

6. Chapter 20 DNA Cloning
Chopping up the whole genome of an organism produces many DNA fragments containing many genes. Often, the researcher will save all of them, either in bacteria or in viruses. These collections of bacterial clones are called genomic libraries.

7. Chapter 20 DNA Probe Hybridization
Samples of DNA

8. Chapter 20 DNA Probe Hybridization
Heat to 95o to denature

9. Chapter 20 DNA Probe Hybridization Introduce radioactive probe

10. Chapter 20 DNA Probe Hybridization
Allow to cool

11. Chapter 20 DNA Probe Hybridization Measure radioactivity
√ x x
Measure radioactivity

12. Chapter 20 Polymerase Chain Reaction (PCR)
Polymerase chain reaction (PCR) uses DNA polymerase to clone DNA in vitro. In vitro = in a test tube In vivo = in a living organism

13. Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Human DNA contains lots of noncoding sequences that serve no purpose.
This “junk DNA” often repeats over and over.
No two people (except identical twins) have exactly the same repeats.

14. gene A xxxxx gene B yyy gene C Bob’s chromosome:
Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Bill’s chromosome:
gene A xxxxx gene B yyy gene C
Bob’s chromosome:
gene A xx gene B yyyyy gene C

15. DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Chapter 20 Gel Electrophoresis
DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Bill’s chromosome:
gene A xxxxx gene B yyy gene C
Bob’s chromosome:
gene A xx gene B yyyyy gene C

16. DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Chapter 20 Gel Electrophoresis
DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Bill’s chromosome:
gene A xxxxx gene B yyy gene C
Bob’s chromosome:
gene A xx gene B yyyyy gene C

17. DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Chapter 20 Gel Electrophoresis
DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Bill’s chromosome:
gene A xxxxx gene B yyy gene C
Bob’s chromosome:
gene A xx gene B yyyyy gene C

18. DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Chapter 20 Gel Electrophoresis
DNA Fingerprinting Restriction enzymes cut DNA at specific places.
Bill’s chromosome:
Bob’s chromosome:

19. Chapter 20 Gel Electrophoresis
DNA Fingerprinting Longer fragments travel more slowly through the gel.
Bill’s chromosome:
Bob’s chromosome:

20. Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Bill: Bob:

21. Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Bill: Bob:

22. Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Bill: Bob:

23. Chapter 20 Gel Electrophoresis
DNA Fingerprinting
Bill: Bob:

24. Chapter 20 Gel Electrophoresis

25. Do DNA fingerprinting on an entire genome.
Chapter 20 Southern Blot
The southern blot
Do DNA fingerprinting on an entire genome.
Blot the DNA from the gel to paper with an alkaline solution. This denatures the DNA.
Hybridize with a radioactive probe.

26. Chapter 20 Southern Blot
The southern blot

27. Chapter 20 Southern Blot
The southern blot

28. Chapter 20 DNA Sequencing
Synthesize DNA 3’ → 5’
Stop synthesis at random
Dideoxyribonuceotides with dye
Separate fragments by length
Read the colors of the lengths.

29. Chapter 20 DNA Sequencing
Synthesize DNA 3’ → 5’

30. Chapter 20 DNA Sequencing
Synthesize DNA 3’ → 5’
5-AGTACCTG-3

31. Chapter 20 DNA Sequencing
Synthesize DNA 3’ → 5’
5-AGTACCTG-3
3-TCATGGAC-5

32. Chapter 20 DNA Sequencing Stop the process with didexoyribonucleotides
5-AGTACCTG-3
3-TCATGGAC-5

33. Chapter 20 DNA Sequencing Stop the process with didexoyribonucleotides
5-AGTACCTG-3
3-TCATG

34. 5-AGTACCTG-3 A T C G Add a different dye to each didexoyribonucleotide
Chapter 20 DNA Sequencing
Add a different dye to each didexoyribonucleotide
5-AGTACCTG-3
A T C G

35. Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
A T C G

36. Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG

37. Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG
3-T

38. 5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC
Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG
3-T
3-TCATGGAC

39. 5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA
Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG
3-T
3-TCATGGAC
3-TCA

40. 5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA 3-TCATG
Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG
3-T
3-TCATGGAC
3-TCA
3-TCATG

41. 5-AGTACCTG-3 3-TCATGG 3-T 3-TCATGGAC 3-TCA 3-TCATG 3-TCA T
Chapter 20 DNA Sequencing
Stop the synthesis at random times with different dyes
5-AGTACCTG-3
3-TCATGG
3-T
3-TCATGGAC
3-TCA
3-TCATG
3-TCA T

42. Chapter 20 DNA Sequencing Arrange the fragments by length
5-AGTACCTG-3
3-TCATGG
3-T
3-TCATGGAC
3-TCA
3-TCATG
3-TCA T

43. Chapter 20 DNA Sequencing Arrange the fragments by length
3-TC
3-TCA
3-TCA T
3-TCATG
3-TCATGG
3-TCATGGA
3-TCATGGAC

44. Chapter 20 DNA Sequencing
Read the colors
3-T
3-TC
3-TCA
3-TCA T
3-TCATG
3-TCATGG
3-TCATGGA
3-TCATGGAC

45. Chapter 20 DNA Sequencing

46. Chapter 20 DNA Sequencing

47. Chapter 20 DNA Technology and Genomics
RFLPs (“RIF-lips”), or restriction fragment length polymorphisms, are differences in homologous chromosomes that give different length restriction fragments. Chromosome walking means finding where fragments of DNA overlapped in the genome.

48. Chapter 20 DNA Technology and Genomics
Genomics is the systematic study of entire genomes. Proteomics is the study of all the proteins encoded by a genome. Single nucleotide polymorphisms (SNPs) are useful markers for studying variation.

49. Chapter 20 DNA Technology and Genomics
Uses of DNA Technology: Testing for genetic diseases Large scale production of drugs Gene therapy Forensics Genetic engineering. .