Frontiers of Biotechnology

1. Frontiers of Biotechnology

2. Manipulating DNA • Scientists use several techniques to manipulate DNA

3. Restriction Enzymes cut DNA • Why cut DNA? • To study specific genes instead of ALL the genes on a chromosome • Restriction enzymes act as molecular scissors • Recognize specific sequences • Some leave “blunt ends” • Some leave “sticky ends”

4. Restriction Maps show the lengths of DNA fragments • Gel Electrophoresis: a technique that uses an electrical field within a gel to separate molecules by their size • DNA is negatively charged and moves toward the positive pole when the electrical field is applied • Smallest DNA fragments move the fastest • A pattern of bands is formed

5. Gel Electrophoresis

6. Polymerase Chain reaction • PCR:technique that produces millions of copies of a specific DNA sequence in just a few hours • Invented by Kary Mullis in 1983

7. PCR • Uses: • DNA to be copied • DNA polymerase • Plenty of nucleotides A, T, C, and G • Two primers • 3 Step Process: • Separating • Binding • Copying

8. RFLPs • Restriction Fragment Length Polymorphisms • No two individuals have the same genetic material except identical twins • Restriction enzymes cut at different places, depending on the DNA sequence • The lengths of DNA restriction fragments are different between two individuals

9. DNA fingerprinting • A DNA fingerprint is a type of restriction map • Representation of parts of a individual’s DNA that can be used to identify a person at the molecular level • Focuses on noncoding regions of DNA, or DNA sequences outside genes

10. DNA Fingerprinting • DNA sample from: • Blood • Semen • Bone • Hair • …Useful in forensics!

11. DNA fingerprinting is used for identification • DNA fingerprints and probability • Compare at least 5 regions of the genome

12. Genetic Engineering • Entire organisms can be cloned • Clone: genetically identical copy of a gene or of an organism • New genes can be added to an organism’s DNA

13. 4 Basic Steps to Genetic Engineering • 1. Cutting DNA • 2. Making recombinant DNA • 3. Cloning • 4. Screening

14. Step 1: Cutting DNA • The DNA from the original organism containing the gene of interest is cut by restriction enzymes • Restriction Enzymes: bacterial enzymes that destroys foreign DNA molecules by cutting them at specific sites

15. Step 1: Cutting DNA • Vector: Any agent, such as a plasmid, that carries the gene of interest into another cell • Plasmid: A circular DNA molecule that is usually found in bacteria and that can replicate independent of the main chromosome

16. Recombinant DNA • DNA molecules that are artificially created • HOW????? • Created by combining DNA from different sources

17. Example: Insulin • A protein hormone that controls sugar metabolism • Diabetics cannot produce enough • Must take doses of insulin daily • Before genetic engineering, insulin was extracted from the pancreases of slaughtered cows and pigs and then purified • Today the human insulin gene is transferred to bacteria through genetic engineering • Because the genetic code is universal, bacteria can transcribe and translate the human insulin gene

18. Step 2: Making Recombinant DNA • DNA fragments from the gene of interest are combined with the DNA fragments from the vector • DNA ligase: an enzyme that bonds the DNA fragments together • The host cell then takes up the recombinant DNA

19. Step 3: Cloning • Gene Cloning: many copies of the gene of interest are made each time the host cell reproduces • Remember: bacteria reproduce by binary fission, producing identical offspring with the plasmid DNA!

20. Step 4: Screening • Cells that have received the particular gene are separated from the cells that did not take up the vector with the gene of interest • The cells can transcribe and translate the gene of interest to make the protein coded for the gene

21. Confirmation of a Cloned Gene • Southern Blot: a technique used to test for the presence of a specific gene

22. Northern Blot • Similar to a Southern Blot • Uses RNA instead of DNA

23. Genetic Engineering produces organisms with new traits

24. Selective Breeding • Allowing only those animals with desired characteristics to produce the next generation • Horses, cats, farm animals, crops

25. Hybridization • Crossing dissimilar individuals to bring together the best of both organisms • Hybrids: the individuals produced from such crosses • For example, a disease resistant plant and the food producing capacity of another

26. Inbreeding • The continued breeding of individuals with similar characteristics • Often seen in dogs • Retains characteristics but has risks • Genetically similar individuals could bring together two recessive alleles for a genetic defect

27. Today…Genetic Engineering

28. Genetically Engineered Crops • More tolerant to drought • Plants that can adapt to different soils, climates, and environmental stresses

29. Genetically Engineered Crops • Resistant to biodegradable weedkillerGlyphosate (kills weeds but now doesn’t kill the crop) • Resistant to insects (gene injures the gut of chewing insects)-therefore plant doesn’t need to be sprayed with pesticides

30. More Nutritious Crops • Improve the nutritious value of many crops • Asia: rice is a staple food • Low in iron an beta carotene • Iron deficient and poor vision • Genetic engineers have added genes to rice from other plants to overcome this deficiency

31. Potential Problems to GM Crops • Concern that some weeds will become resistant to the weed killer Glyphosate • New weed-control alternatives will have to be implemented

32. Potential Problems to GM Crops • Nutritional value has been increased in many crops • Crops must be tested to make sure consumers are not allergic to the GM product

33. Gene Technology: Animal Farming • Farmers added growth hormones to the diet of cows to increase milk production • Growth hormone was extracted from the brains of dead cows • The hormone was introduced into bacteria and added as a supplement to a cow’s diet

34. Transgenic Animals • Animals that have foreign DNA in their cells • Human genes have been added to farm animals in order to get the farm animals to produce human proteins in their milk

35. Transgenic Animals • This is complex and cannot be made by bacteria through gene technology • Human proteins are extracted from the animal’s milk and sold for pharmaceutical purposes • Cloning animals: creating herds of identical animals that can make medically useful proteins

36. Cloning from Adult Animals • The intact nucleus of an embryonic or fetal cell (whose DNA has been recombined with a human gene) is placed into an egg whose nucleus has been removed • The egg with the new nucleus is put in the uterus of a surrogate, or substitute mother and allowed to develop

37. Cloning from Adult Animals • 1997 Ian Wilmut first successful cloning using differentiated cells from an adult animal • Dolly the sheep

38. Cloning from Adult Animals • Differentiated cells: cells that have become specialized to become specific cell types • Scientists had thought that embryonic or fetal cells were the only way…wrong!

39. Cloning from Adult Animals • Mammary cells from one sheep were fused with egg cells without nuclei form a different sheep • The fused cells divided to form embryos, implanted into surrogate mothers • Only one survived the cloning process • Dolly, identical to the sheep that provided the mammary cell

41. Problems with Cloning • Only a few of the cloned offspring survive for long • Many become fatally oversized • Problems in development

42. Genomic Imprinting • The right combination of genes are turned “on” and “off” during early development • The egg takes years to develop the genomic imprint • In cloning, the egg divides within minutes

43. Genomic Imprinting • Reprogramming is not possible in such a short time • Critical errors in development can occur • Because of these technical problems and ethical problems, cloning humans is illegal in most countries

44. Concerns about genetic engineering • Ethical? • GM crops • Not enough research had been done to see if added genes might cause allergic reactions or have other unknown side effects • Interbreeding with natural plants…what does it mean?

45. Genomics involves the study of genes, gene functions, and entire genomes • Genomics: The study of genomes, which can include the sequencing of all of an organism’s DNA • Gene sequencing: determining the order of DNA nucleotides in genes or in genomes

46. The Geography of the Genome • Only 1-1.5% of the human genome codes for proteins • Each human cell contains about 6 feet of DNA • Less than 1 inch are exons

47. The Geography of the Genome • Human cells contain about 25,000 genes (scientists had expected 120,000!) • Only 2x the number of genes in a fruit fly! • Many human genes are identical to those of other species • All humans are genetically close (DNA of any 2 people is 99.9% identical)

48. The human genome project • Our genome is relatively small! 3 billion base pairs, but only between 30,000-40,000 genes • Project started in 1990 with 2 main goals: • Map and sequence all of the DNA base pairs of the human chromosomes (accomplished in 2003) • Identify all of the genes within the sequence (still be worked on)

49. The human microbiome project • 200 scientists at 80 institutions sequenced the genetic material of bacteria taken from nearly 250 healthy people • As many as a thousand bacterial strains on each person. • Each person’s collection of microbes, the “microbiome”, was unique

50. Technology allows the study and comparison of both genes and proteins • Bioinformatics: the use of computer databases to organize and analyze biological data • DNA microarrays: tools that allow scientists to study many genes, and their expression at once; a small chip dotted with the genes being studied • Proteomics: the study and comparison of all the proteins that result from an organism’s genome