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Chapter 12

Chapter 12. Genetic Enginneering. "In Science the credit goes to the man who convinces the world, not to the man to whom the idea first occurs." – Sir Francis Darwin (1848-1925, English botanist, son of Charles Darwin), In: Eugenetics Review (April 1914, "Francis Galton").

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Chapter 12

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  1. Chapter 12 Genetic Enginneering

  2. "In Science the credit goes to the man who convinces the world, not to the man to whom the idea first occurs."– Sir Francis Darwin (1848-1925, English botanist, son of Charles Darwin), In: Eugenetics Review (April 1914, "Francis Galton")

  3. At the International Genetics Meeting. • An American reports:"We crossed chickens with cows. The new breed simultaneously produces milk, meat and eggs.“ • A Frenchman reports:"We succeeded in cross-breeding flies and bees. The hybrid flies over the trash fields and produces honey.“ • A Russian reports:"And we crossed a melon with cockroaches. When you cut this melon, seeds run away by themselves."

  4. DNA Sequencing

  5. Genes can be cloned in Recombinant Plasmids

  6. b

  7. Genes can be stored in genomic Libraries

  8. Use of Reverse Transcriptase

  9. Nuc. Acid Probes are used to identify genes of interest

  10. s

  11. Gel Electrophoresis apparati

  12. Bad gels

  13. Southern Blots • Figure 5.35 from Hart & Jones, 1998 (page 207) • First, you run a "normal" DNA gel • (usually agarose, but it doesn't HAVE to be). • then you hybridize the DNA onto a membrane

  14. Next, this membrane is soaked in a mixture containing radioactively labelled probe, which should hybridize

  15. The filter is washed and exposed to radioactive film.

  16. RFLPs • restriction enzyme-generated fragments of different length • molecular markers used in creating genetic maps of chromosomes. • relies on hybridization between a probe and homologous DNA segments within a genome.

  17. Polymorphism means the existence of many different forms in which a region of DNA can have different lengths on different homologs or in different individuals.

  18. RFLP analysis

  19. Although two individuals of the same species have almost identical genomes, they will always differ at a few nucleotides. • Some of these differences will produce new restriction sites (or remove them), and the banding pattern seen on a genomic Southern will thus be affected. • For any given probe (or gene), it is often possible to test different restriction enzymes until one which gives a pattern difference between two individuals is found, that is, a RFLP. • The less related the individuals, the more divergent their DNA sequences are and the more likely you are to find a RFLP.

  20. a semen stain left on the victim's clothing (EVIDENCE #1); semen removed from the vagina of the rape victim (EVIDENCE #2); the DNA of the victim herself (VICTIM) to be sure that the DNA didn't come from her cells; DNA from two suspects a set of DNA fragments of known and decreasing length (MARKER).. the cells of a previously-tested person to be sure the probes are performing properly (CONTROL).

  21. thermal cycler for PCR

  22. Chromosome walking is a method in genetics for identifying and sequencing long parts of a DNA strand,. The basic technique is as follows: A primer that matching the beginning of the DNA to sequence is used to synthesize a short DNA strand complementary to the unknown sequence, starting with the primer (see PCR). The new short DNA strand is sequenced using the chain termination method. The end of the sequenced strand is used as a primer for the next part of the long DNA sequence.

  23. Linkage Mapping Allows to tell where they are using recombination frequencies • DNA Sequencing • Physical Map ( order of fragments) • Chromosome walking

  24. Transposons • Insertion Sequences or IS elements simplest mobile element. • consist of a fairly short (700 - 1500 bp) DNA segment flanked by a 10 - 40 bp inverted repeat sequence. • The segment codes for the protein (transposase) that catalyses the transposition event:

  25. Composite transposons are DNA segments flanked by an IS element at either end. • In other words, instead of each IS element moving independently, they now act in concert and move together along with the intervening DNA.

  26. Practical Applications 1. Medical Applications • Diagnosis of Diseases

  27. Human Gene Therapy

  28. 2. Pharmaceutical Products Vaccines, Growth hormones, insulin, etc… 3. Forensics Evidence DNA fingerprints from a murder case. This autoradiograph shows that DNA in blood from the defendant's clothes matches the DNA fingerprint of the victim but differs from the DNA fingerprint of the defendant. This is evidence that the blood on the defendant's clothes came from the victim, not the defendant.

  29. 4. Environmental Cleanups -Oil spills -Mining Bacteria being sprayed onto soil prepared for Bio-Remediation

  30. 5. Agricultural Applications Pharm” animals.  These transgenic sheep carry a gene for a human blood protein, which they secrete In their milk. This protein inhibits an enzyme that contributes to lung damage in patients with cystic fibrosis and some other chronic respiratory diseases. Easily purified from the sheep′s milk, the protein is currently under evaluation as a treatment for cystic fibrosis.

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