1. Chapter 16 Genetic Engineering

2. Beginnings Cholesterol- good, necessary, naturally made by our body. Can be bad though –Cholesterolemia- too much cholesterol made causing early heart attacks in teens Gene therapy- add altered or normal genes to a cell to correct a defect or to boost the natural systems

3. Gene therapy

4. Beginnings For nearly 3 billion years, mutation, crossing-over, random gene-mixing at fertilization, and hybridizations have contributed to the diversity of life on Earth Today we can “engineer” genetic changes through recombinant DNA technology –DNA splicing using bacteria –Genetic engineering has great potential in agriculture, medicine, industry –Ecological, social, ethical questions and concerns

5. 15.1 Tools necessary Restriction enzymes, modification enzymes, cloning vectors, reverse transcriptase Restriction enzymes-bacteria usually possess enzymes whose function is to cut apart foreign DNA –Cuts specific base sequences –Wide variety makes it possible to study the genome of a particular species

6. Modification enzymes The “sticky” ends from the sections cut by restriction enzymes can be used to pair up with other DNA fragments These fragments that stick together are then treated with DNA ligase to splice the DNA together to make a recombinant DNA molecule

7. Recombinant DNA

8. Cloning vectors Amplify DNA Plasmids are circular DNA molecules in bacteria that carry a few genes separate from the main chromosome When plasmid is replicated any foreign DNA that the bacteria has come into contact with is also replicated producing a DNA clone Modified plasmids are capable of accepting, replicating, and delivering DNA to another host cell are called cloning vectors

9. Plasmids

10. Reverse transcriptase Even after a gene has been isolated and amplified, it may not be translated because the introns (noncoding regions) are still present Researchers minimize this problem by using cDNA (complementary DNA) –Made using reverse transcription. This makes DNA backwards from RNA –cDNA can be inserted into a plasmid –Used by viruses

11. Viral gene therapy

12. 16.2 PCR Polymerase chain reaction- used to make millions of copies of cDNA –Primers- short nucleotide segments made in a lab –Recognized by DNA polymerase at the start tags dictated by computers –Researchers mix primers, DNA polymerase, cellular DNA from the org., free nucleotides –Temperature cycles cause the DNA to unwind –Primers become positioned on the exposed bases to form new copies of the original –Each round doubles the amount of DNA molecules to produce billions of copies –http://members.aol.com/BearFlag45/Biology1A/Lectur eNotes/lec24.html

13. 16.4 DNA sequencing Current labs use automated DNA sequencing to determine the unknown sequences in a sample –Uses 8 bases to build DNA molecules –The bases are modified fluoresce in a laser –When a modified base is incorporated, DNA synthesis is halted producing tagged fragments Tagged sequences are identified by laser and then copied using PCR The short segments are then assembled to reveal the entire sequence

14. DNA sequencing

15. 16.5 Isolating Genes Using DNA from bacterial cloning create a library of gene fragments Library may consist of entire genome or cDNA which is free of introns DNA probes- short sequences of DNA assembled from radioactive nucleotides- can be paired up with gene parts to be studies –This nucleic acid hybriduzation technique can be used with other procedures to select cells and their DNA

16. Screening for genes Grow the bacteria on plates Cover with nylon filter and remove some cells Place filter in a solution to disrupt cells but leave DNA on the filter Add radioactively-labeled probe DNA to filter where it binds to DNA fragments of complementary sequence Expose the filter to x-ray film to locate the gene of interest which will be the same as the cells in the dish

17. 16.6 Using the Genetic scripts –Bacteria used to produce insulin and blood- clotting factors –Bioremediation- using genetically engineered bacteria to clean up oil spills –Helps us develop counter attacks to mutated pathogens

18. 16.7 Designer plants Regenerating plants from cultured cells –Researchers are searching the world for seed of wild potatoes, corn, etc.. –The worry is that there is too little variation in the few strains used for crops –Many species can be regenerated from cultured cells –Useful mutations, such as resistance to toxin, can be identified

19. Transferring genes Early experiments showed that a plasmid from a bacteria that normally causes tumors in plants could be modified by replacing the tumor gene with desirable genes Modified bacteria have been injected into plant cells where they express their “foreign” genes –Genetically modified foods can increase food production of develop greater resistance to pests –Can be used to produce human hemoglobin, melanin, plastics –BT corn uses Bacillus therangiensis DNA

20. BT Corn & Genetic modification

21. 16.8 Gene transfer in animals Supermice- 1982- rat gene for somatotropin was introduced into mice eggs. The mice that expressed the gene grew larger than their littermates Farm animals may be used to produce TPA for treating heart attacks, goats used to produce CFTR which is used to treat cystic fibrosis Cloning of animals could lead to disease-resistant types Dolly

22. Using the human genome 1980’s began interest in mapping human genome, 2003 completed Dependent on technology discussed Gain insights into genetic disorders, gene therapy Gain insight into the evolutionary relationships between organisms http://www.ornl.gov/sci/techresources/Human_Ge nome/home.shtml

23. 16.9 Safety Genetically engineered bacteria have “fail- safe” genes to ensure that they do not survive outside the lab Public concerns over “natural” vs “engineered” crops May endanger human lives or natural plants

24. 16.10 Biotechnology Microarrays or gene chips Gene therapy has been successful in a trial against SCID-X1 (severe combined immunodeficiency disorder- X-linked)- bubble-boys Which disorders will receive attention Eugenics- genetic racism- GATTACA Xenotransplantation- transplanting organs from one species into another Pigs can be engineered to lack certain genes that would cause rejection problems in humans. Problems? Should we alter the DNA of organisms? Some say we have through selection. How much is too much?